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Divergent Evolution of Lanthipeptide Stereochemistry
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  • Raymond Sarksian
    Raymond Sarksian
    Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States
    More by Raymond Sarksian
    Orcidhttps://orcid.org/0000-0001-6940-6699
  • Wilfred A. van der Donk*
    Wilfred A. van der Donk
    Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States
    Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States
    *Email: [email protected]. Tel: 217 244 5360.
    More by Wilfred A. van der Donk
    Orcidhttps://orcid.org/0000-0002-5467-7071
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ACS Chemical Biology

Cite this: ACS Chem. Biol. 2022, 17, 9, 2551–2558
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https://doi.org/10.1021/acschembio.2c00492
Published August 24, 2022

Copyright © 2022 The Authors. Published by American Chemical Society. This publication is licensed under

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Abstract

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The three-dimensional structure of natural products is critical for their biological activities and, as such, enzymes have evolved that specifically generate active stereoisomers. Lanthipeptides are post-translationally modified peptidic natural products that contain macrocyclic thioethers featuring lanthionine (Lan) and/or methyllanthionine (MeLan) residues with defined stereochemistry. In this report, we compare two class I lanthipeptide biosynthetic gene clusters (BGCs), coi and olv, that represent two families of lanthipeptide gene clusters found in Actinobacteria. The precursor peptides and BGCs are quite similar with genes encoding a dehydratase, cyclase, and methyltransferase (MT). We illustrate that the precursor peptide CoiA1 is converted by these enzymes into a polymacrocyclic product, mCoiA1, that contains an analogous ring pattern to the previously characterized post-translationally modified OlvA peptide (mOlvA). However, a clear distinction between the two BGCs is an additional Thr-glutamyl lyase (GL) domain that is fused to the MT, CoiSA, which results in divergence of the product stereochemistry for the coi BGC. Two out of three MeLan rings of mCoiA1 contain different stereochemistry than the corresponding residues in mOlvA, with the most notable difference being a rare d-allo-l-MeLan residue, the formation of which is guided by CoiSA. This study illustrates how nature utilizes a distinct GL to control natural product stereochemistry in lanthipeptide biosynthesis.

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Introduction

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Natural products typically recognize their targets with exquisite affinity and selectivity. (1) In the course of their evolution, the structures of these molecules have been optimized to bind to the usually chiral environments of biological targets. This high level of recognition is often achieved by the rich stereochemistry of natural products, which have made them privileged ligands. (1) In this study, we report the discovery of two lanthipeptides that have very similar ring patterns but in which the stereochemistry of two of the three macrocycles differs. We show that the acquisition of one new enzyme has driven the divergence of the two compound groups and that this new activity has resulted in coevolution of other enzymes in the pathway.
Lanthipeptides represent one of the largest classes of ribosomally synthesized and posttranslationally modified peptides (RiPPs). (2−4) They exhibit a wide range of activities, including antimicrobial, antiviral, morphogenetic, and antifungal, (3,5−10) and are defined by the presence of lanthionine (Lan) or methyllanthionine (MeLan) residues (Figure 1). Maturation of lanthipeptides features the dehydration of Ser/Thr residues followed by subsequent Michael-type addition of a Cys thiol onto the dehydroamino acids to form (Me)Lan (Figure 1). (2−4)

Figure 1

Figure 1. (A) Maturation of lanthipeptides proceeds through dehydration of Ser/Thr residues to generate the corresponding dehydroamino acids dehydroalanine and dehydrobutyrine (Dha, Dhb). Cyclization of Cys side chains onto Dha/Dhb yields (Me)Lan. (B) Four possible MeLan diastereomers can be generated as shown.

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For class I lanthipeptides, a lanthipeptide dehydratase LanB first catalyzes the transesterification of a glutamate group from glutamyl-tRNAGlu to the side chain of Ser/Thr. (11−14) An elimination reaction of the glutamylated intermediate generates dehydroalanine (Dha) from Ser or dehydrobutyrine (Dhb) from Thr (Figure 1A). A LanC cyclase then catalyzes the formation of thioether rings by the addition of a Cys thiol to the dehydroamino acid intermediates. (15,16)
The stereochemical configuration of (Me)Lan residues has been shown to be important for the biological activities of lanthipeptides. (17,18) Three MeLan diastereomers have been discovered to date, (2S,3S,6R)-, (2R,3R,6R)-, and (2S,3R,6R)-MeLan, hereafter referred to as dl-, ll-, and d-allo-l-MeLan (Figure 1B). (4,19−25) Both dl- and ll-MeLan are believed to form through the anti-elimination of Thr residues to yield (Z)-Dhb residues followed by an anti-addition of Cys across the (Z)-Dhb. (4) Facial selectivity of the cyclization event dictates whether dl- or ll-MeLan is formed from the (Z)-Dhb. (20)d-allo-l-MeLan was recently reported for the morphogenetic class I lanthipeptide SapT. (24)d-allo-l-MeLan is thought to be formed through syn-elimination of glutamylated Thr residues followed by subsequent anti-addition of Cys across the (E)-Dhb. (24) The SapT biosynthetic gene cluster (BGC) features a split dehydratase made up of SptBa and SptBb that carries out dehydration. SptBa catalyzes glutamylation of Ser/Thr residues, and SptBb is a glutamyl lyase (GL) that catalyzes syn-elimination. (24)
In this study, we compare the coi BGC from Streptomyces coelicolor A3(2) with the olv BGC from Streptomyces olivaceus NRRL B-3009 (Figure 2A). These BGCs are representative examples of two groups of gene clusters in Actinobacteria (Figure 2A), with only the olv BGC previously investigated in depth. (26) Both BGCs encode a canonical class I dehydratase (CoiB and OlvB) and cyclase (CoiC and OlvC), and an O-methyltransferase (MT) that is the most widespread auxiliary enzyme in class I lanthipeptide BGCs. (26−28) The sequences of their precursor peptides are also quite similar (Figure 2B). The ring pattern and stereochemistry of the (Me)Lan residues of the olv product have been determined by nuclear magnetic resonance (NMR) spectroscopy and comparison with synthetic standards. (26) The coi BGC mainly differs from the olv BGC in that the MT CoiSA has a fused GL domain and encodes a protein of unknown function CoiH. (24,26)

Figure 2

Figure 2. (A) Comparison of coi and olv class I lanthipeptide BGCs. The coi BGC encodes an additional GL and protein of unknown function when compared to the olv BGC. (B) Sequence alignment of the precursor peptides encoded in both BGCs. Residues that are known to be modified in OlvA are colored. (C) Sequence alignment of GL domains illustrates that the additional GL domain in CoiSA is similar to SptBb.

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The presence of two GLs in the coi BGC is unusual. The GL domains in both CoiB and CoiSA contain conserved Arg residues that are important for the recognition of the γ-carboxylate of the glutamylated peptide intermediate in lanthipeptide dehydratases (Figure 2C). (11−14) However, only CoiB contains the catalytic His base and Arg residue that are critical for glutamate elimination activity in canonical anti-GLs (Figures 2C and S1). (11−14) The GL domain in CoiSA in contrast contains similar putative active site residues as SptBb. (24)
Here, we investigate the regioselectivity of the two distinct GL domains in CoiB and CoiSA (Figure 2). Both enzymes catalyze glutamate elimination; however, they are proposed to generate two different Dhb isomers during the maturation of CoiA1. (24) We demonstrate that the 3-fold dehydrated and cyclized product, mCoiA1, contains three different MeLan diastereomers in a ring pattern that is very similar to the olv product. However, elucidation of the stereochemical configuration for each MeLan residue showed that two of the three residues have different stereochemistry compared to that found in the olv product. Furthermore, our data show that CoiC catalyzes cyclization with both (Z)- and (E)-Dhb residues but only when these isomers are at their native location, suggesting coevolution of the cyclase with the product stereochemistry. These findings provide intriguing insights into the divergent evolution of two widespread natural products by acquisition (or deletion) of a GL that results in different stereochemistry.

Results

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Heterologous Production and Characterization of mCoiA1

The coi cluster is present in the genome of one of the most widely studied strains of Streptomyces, S. coelicolor A3(2), (29−32) but its product has not been previously observed despite considerable genome mining studies. (33−37) Therefore, in this study, we used heterologous expression to investigate its product. CoiA1 was previously shown to undergo 3-fold dehydration and cyclization to yield mCoiA1 when coexpressed with CoiB, the elimination domain of CoiSA (CoiSA(ED)), and CoiC in Escherichia coli. (24) In this work, the coexpressed product was isolated and treated with endoproteinase GluC (Figure 3). High-resolution mass spectrometry (HRMS) analysis confirmed that the product was dehydrated three times (Figure 3A). Tandem MS of the GluC-digested peptide suggests that it contains an N-terminal MeLan ring and two C-terminal overlapping MeLan rings (Figure 3B, Table S3). This ring pattern is similar to that determined by tandem MS and NMR spectroscopy for the olv product, (26) consistent with the conservation of the positions of the Ser/Thr and Cys residues in the CoiA1 and OlvA precursor peptides (Figure 2B).

Figure 3

Figure 3. Liquid chromatography-MS (LC-MS) analysis of the GluC digestion product of mCoiA1 (mCoiA1GluC). (A) High-resolution electrospray ionization-MS (ESI-MS) analysis. (B) Tandem ESI-MS analysis. Fragmentation results are consistent with an N-terminal MeLan and two C-terminal overlapping MeLan.

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Glutamate Elimination Activity

Next, we performed experiments to gain insight into the elimination activity of both CoiB and CoiSA(ED). Coexpression of CoiA1 with CoiB and CoiC led to at most one dehydration along with intermediates that were glutamylated once or twice (Figure 4). This finding suggests that the C-terminal lyase domain of CoiB was only able to perform one elimination of the Ser/Thr residues that were glutamylated by the N-terminal domain of CoiB. In contrast, when CoiA1 was coexpressed with CoiSA(ED), CoiC, and the CoiB-H994A mutant, in which the lyase activity of the C-terminal domain of CoiB was inactivated but the glutamylation activity was retained, (12,14) a 3-fold dehydrated product was observed (Figure S2). Thus, CoiSA(ED) was able to eliminate glutamate at all three Thr residues.

Figure 4

Figure 4. Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS analysis of coexpression experiments in E. coli. Coexpressed proteins are listed on each panel. Asterisks indicate deamination products that are commonly generated in MALDI-TOF mass spectra at these masses.

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An N-ethylmaleimide (NEM) alkylation assay (38) to test for the presence of free Cys suggested that the product peptide of the latter experiment was not fully cyclized (Figure S3), implying that the 3-fold dehydrated peptide generated by CoiSA(ED) is not a competent substrate for CoiC. These results show that both CoiB and CoiSA(ED) are required for correct dehydration and cyclization of CoiA1. Finally, we investigated whether CoiC is necessary to obtain a 3-fold dehydrated product. In the case of some lanthipeptides such as microbisporicin A1, (13) the lanthipeptide cyclase is required to obtain full dehydration since select (Me)Lan rings must form prior to the next dehydration event. Coexpression of CoiA1 with CoiB and CoiSA(ED) resulted in a 3-fold dehydrated product (Figure 4). Therefore, CoiC and the formation of (Me)Lan rings are not necessary for combined CoiB and CoiSA(ED) activity.

Stereochemical Analysis of MeLan Residues

mCoiA1 was previously demonstrated to contain dl-, ll-, and d-allo-l-MeLan residues (Figure 5A). (24) Assignment of the stereochemistry to specific rings was not reported. Since mCoiA1 only contains MeLan rings, Thr residues involved in ring formation were individually mutated to Ser residues to determine the stereochemistry for each MeLan ring. (25) If successful, this approach would remove a single MeLan residue (by conversion to a Lan residue) in each variant and thus allow assignment of stereochemistry to that MeLan.

Figure 5

Figure 5. Gas chromatography-MS (GC-MS) analysis of derivatized MeLan residues (m/z = 379) obtained from mCoiA1 mutant peptides using a chiral stationary phase. Coinjections with stereochemically pure derivatized dl- and ll-MeLan confirm the presence of dl-MeLan in mCoiA1-T43S and ll-MeLan in mCoiA1-T50S. (A) Derivatized MeLan from WT mCoiA1 (top), synthetic standards, and coinjections of the products of mCoiA1 with the synthetic standards. (B) Derivatized MeLan from mCoiA1-T43S (top), synthetic standards, and coinjections of the products of mCoiA1-T43S with the synthetic standards. (C) Derivatized MeLan from mCoiA1-T50S (top), synthetic standards, and coinjections of the products of mCoiA1-T50S with the synthetic standards. (D) Structures of derivatized dl- and ll-MeLan standards.

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CoiA1-T43S, CoiA1-T50S, and CoiA1-T57S variants were generated by site-directed mutagenesis and coexpressed with CoiB, CoiSA(ED), and CoiC followed by isolation of the products by Ni-affinity chromatography. mCoiA1-T43S and mCoiA1-T50S underwent 3-fold dehydration and were unreactive toward NEM, suggesting that the peptides were cyclized (Figure S4). HRMS and tandem MS confirmed that the cyclization patterns of these two mutant peptides were not altered from WT mCoiA1 (Figures S5 and S6). In contrast, mCoiA1-T57S was obtained in multiple dehydration states and was not fully processed (Figure S4). Hence, only mCoiA1-T43S and -T50S were hydrolyzed in acid and the resulting amino acids were derivatized for stereochemical analysis.
Stereochemical analysis was performed by gas chromatography-mass spectrometry (GC-MS) on a chiral stationary phase and comparison to authentic dl- and ll-MeLan standards. (25,39,40) mCoiA1-T43S contained two MeLan, as anticipated (Figure 5B). Coinjections with stereochemically pure dl- and ll-MeLan confirmed the presence of dl-MeLan and absence of ll-MeLan (Figure 5B). Therefore, WT mCoiA1 must contain an N-terminal ll-MeLan ring derived from Thr43. Using the same approach, mCoiA1-T50S also revealed peaks in the GC-MS corresponding to two MeLan (Figure 5C). Coinjections confirmed one of the MeLan peaks to consist of MeLan with the ll-configuration and dl-MeLan was absent in the sample. By the same logic, WT mCoiA1 must contain a dl-MeLan ring derived from Thr50.
The second-eluting MeLan peak from mCoiA1-T43S and mCoiA1-T50S that did not match either dl- or ll-MeLan was anticipated to be the rare allo isomer as previously detected in WT mCoiA1. (24) The two possible allo-MeLan stereoisomers were previously shown to be inseparable by GC-MS. (24) Hydrolysis of mCoiA1-T43S and mCoiA1-T50S and derivatization of the amino acids with Marfey’s reagent, Nα-(2,4-dinitro-5-fluorophenyl)-l-alaninamide (l-FDAA), followed by comparison to d-allo-d/l-MeLan and l-allo-d/l-MeLan standards (24) confirmed the presence of d-allo-d/l-MeLan in both peptides by liquid chromatography-mass spectrometry (Figure S7). mCoiA1 was next demonstrated to contain d-allo-l-MeLan and not d-allo-d-MeLan by isolation of MeLan and reductive desulfurization, which would form d-Ala from d-allo-d-MeLan or l-Ala from d-allo-l-MeLan (Figure S8). (9) The desulfurization product consisted of l-Ala (Figure S8) confirming the presence of d-allo-l-MeLan. Because this isomer is also seen in both mCoiA1-T43S and mCoiA1-T50S, the MeLan derived from Thr57 in mCoiA1 must be d-allo-l-MeLan. Based on the sequence similarity of CoiSA(ED) with SptBb, the rare d-allo-l-MeLan is generated through the involvement of CoiSA(ED). The poor conversion of CoiA1-T57S suggests that mutation of Thr57 to Ser is not well tolerated by CoiSA(ED) and may imply that some syn-GLs are specific for Thr.

CoiSA(ED) Mutational Analysis

Mutational analysis was next performed on CoiSA(ED) to decipher the importance of putative active site residues. Based on sequence analysis, CoiB is very similar to the nisin dehydratase NisB and related class I lanthipeptide dehydratases that catalyze anti-elimination (Figure S1). (11−14) GLs that catalyze anti-elimination have been well characterized both biochemically and structurally. In contrast, syn-GLs have only recently been discovered, and their putative active sites diverge from anti-GLs (Figures 2C and S1). CoiSA(ED), SptBb, and related homologs contain a highly conserved Lys residue that when mutated in CoiSA(ED) (CoiSA(ED)-K46A) resulted in the accumulation of glutamylated peptides implying its importance for glutamate elimination. (24)
In addition to the differences observed between syn- and anti-GLs, some key sequence similarities are also found. In NisB, Arg784 and Arg786 bind to the γ-carboxylate of the glutamylated peptide intermediate and are critical for elimination activity. (12−14) These residues are also conserved in CoiSA(ED), and Ala mutants were generated to determine the importance of activity. Both CoiSA(ED)-R51A and -R53A mutants were coexpressed with CoiA1, CoiB, and CoiC. For both mutants, a 3-fold dehydrated product was obtained, along with a 2-fold dehydrated product (Figure 6). Therefore, in contrast to NisB, the Arg residues do not seem to be absolutely critical as elimination activity is not severely diminished. Finally, we generated CoiSA(ED)-E89A. This Glu is highly conserved across all class I lanthipeptide GLs (Figure S1; Glu823 in NisB), and based on a calculated structure of SptBb, the residue points toward the putative active site and may play a role in catalysis. (24) Coexpression experiments of CoiA1, CoiB, and CoiC with the CoiSA(ED)-E89A mutant revealed that dehydration activity was altered, but a 3-fold dehydrated peptide was still generated (Figure 6). Thus, Glu89 is also not critical for catalysis by CoiSA(ED).

Figure 6

Figure 6. MALDI-TOF MS analysis of CoiA1 coexpression with CoiB, CoiC, and CoiSA(ED) WT or CoiSA(ED) mutants. Asterisks indicate deamination products that are commonly generated in MALDI-TOF mass spectra at these masses.

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Bioinformatic Analysis and Comparison of anti- and syn-GLs

Two key differences exist between anti- and syn-GLs investigated thus far based on sequence and mutational analysis. A highly conserved Arg residue (Arg826 in NisB) is present in anti-GLs (Figure 2C) that interacts with the carbonyl oxygen of glutamylated Thr to lower the pKa of the α-proton and facilitate elimination. (14) For SptBb, CoiSA(ED), and related homologs, this residue is replaced by a hydrophobic residue (Figure 2C). In addition, SptBb and CoiSA(ED) contain a highly conserved Lys residue that is important for elimination activity that is a Tyr in anti-GLs.
We surveyed all GLs involved in the BGCs of lanthipeptides and related RiPPs using these sequence differences. A sequence-similarity network (SSN) for GLs was previously generated using the tools of the Enzyme Function Initiative (Figure 7). (41,42) In this study, the genomic context of each group was inspected and the GLs in the class I lanthipeptide BGCs were selected for sequence alignments to determine whether they belong to anti-GLs or syn-GLs (Figure 7 and Table S6). This analysis suggested grouping of the GLs into three broad categories. The largest group of GLs is the full-length LanB proteins that are all predicted to catalyze anti-elimination (Figures 7, black, and S1). Both CoiB and OlvB are found within this group. Smaller groups of anti-GLs are part of split LanB systems with dedicated glutamyl transferases and glutamate lyases (Figures 7, blue, and S1). Consistent with a previous study, (24) a significant portion of the network also revealed syn-GLs as (1) part of a split LanB system, (2) fused to a methyltransferase domain such as CoiSA, or (3) present as an additional stand-alone GL domain, with the latter two in BGCs that also contain a full-length dehydratase (Figure 7, purple). All other groups containing more than two members were inspected and found to be part of BGCs of other RiPPs such as thiopeptides (Figure 7, gray, and Table S6). Analysis of GLs within these BGCs suggests that the associated GLs are similar to anti-GLs and are likely generating (Z)-Dhb and/or Dha residues as either intermediates and/or in the final products. Hence, the occurence of syn-GLs seems to be limited in the currently sequenced genomes to class I lanthipeptide BGCs. The SSN also provided the opportunity to assess whether (E)-Dhb and/or allo-MeLan isomers are present in any previously reported class I lanthipeptides for which stereochemistry has generally not been determined. The active site residues of GLs involved in the biosynthesis of these previously characterized family members were analyzed (Figure S9). These GLs were found to be very similar to anti-GLs suggesting that (E)-Dhbs and/or allo-MeLan isomers are likely not present in these lanthipeptides.

Figure 7

Figure 7. SSN analysis of GLs. All colored groups indicate GLs within class I lanthipeptide BGCs. Full-length LanB dehydratases that catalyze anti-elimination are shown in black, and anti-GLs within split LanB systems in blue. Syn-GLs that are part of a split LanB system that are fused to a methyltransferase or that are present as stand-alone proteins are depicted in purple. GLs within BGCs of other nonlanthipeptide or hybrid RiPPs are in gray. A select number of characterized GLs are labeled including the thiopeptide GLs TbtC and LazF. For brief summaries of the biosynthetic genes in each group, see Table S6. The cytoscape file for the SSN is provided as the Supporting Information.

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Discussion

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The overall structure of mCoiA1 in terms of the stereochemical configuration of (Me)Lan residues is the most complex of any lanthipeptide characterized thus far. mCoiA1 contains an N-terminal ll-MeLan ring and overlapping C-terminal dl- and d-allo-l-MeLan rings (Figure 8A). Except for an additional Lan in mOlvA, mCoiA1 and mOlvA contain (Me)Lan rings at equivalent positions. It is interesting to note that the Ser residue that is involved in this additional N-terminal Lan in OlvA is also present in CoiA1 but escapes dehydration. The corresponding Cys residue to generate the Lan ring is missing in CoiA1 and replaced by an Asn residue (Figures 2B and 8A).

Figure 8

Figure 8. (A) Comparison of the structures of GluC-digested mCoiA1 and mOlvA to highlight differences and similarities in ring patterns and stereochemistry. Abu, 2-aminobutyric acid. The N-terminal Lan ring in mOlvA is absent in mCoiA1, although the corresponding Ser residue (gold) is present. (B) Sequence LOGO made using orthologs of the core peptide region of OlvA and CoiA1 identified by BLAST analysis (45) to highlight conserved residues (OlvA total sequences = 43, CoiA1 total sequences = 301).

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Although the ring patterns of mOlvA and mCoiA1 are similar, the stereochemical configurations of two of the three (Me)Lan rings differ. In the C-terminal region of these peptides, mOlvA contains an ll-Lan, whereas mCoiA1 contains a rare d-allo-l-MeLan residue (Figure 8A). A critical distinction between the coi and olv BGCs is the presence of the additional GL domain in CoiSA, which is similar to SptBb that was previously proposed to catalyze syn-elimination of glutamylated Thr to generate (E)-Dhb that leads to the observed d-allo-l-MeLan in SapT. (24) The second deviation in stereochemistry is that mCoiA1 contains an N-terminal ll-MeLan ring where mOlvA contains a dl-MeLan ring at the equivalent position (Figure 8A). Unlike the change in stereochemistry at the C-terminus, which is accounted for by the presence of a gene encoding an additional syn-GL in the BGC, the origin of the change in the stereochemistry of the N-terminal MeLan is less clear. The conformational landscape of lanthipeptides has been shown to be important for the cyclization reaction. (43,44) Therefore, one possibility is that the additional N-terminal ll-Lan ring in mOlvA is formed early during the biosynthetic process and that this ring conformationally biases the peptide toward forming an alternative diastereomer for the adjacent ring. Alternatively, the change in the stereochemistry of the C-terminal MeLan because of the recruitment of a new GL could in turn also influence the stereochemistry of the N-terminal ring if the d-allo-l-MeLan is formed early in the maturation process. Regardless of the molecular explanation of the change in stereochemistry, in all investigated examples, engineered changes in stereochemistry have led to the abolishment of the original bioactivity of the lanthipeptide. (17,18) Therefore, it is likely that the differences in stereochemistry between the products of the widespread coi and olv gene cluster families have functional consequences.
Several examples have been reported wherein RiPP biosynthesis requires an obligate order of post-translational modifications. (3) The investigation of the coi BGC provides another example of high coordination of the post-translational modification reactions. Based on the stereochemistry of the final product, it is likely that CoiB converts Thr43 and Thr50 into (Z)-Dhb residues, which are then the substrates for CoiC-catalyzed cyclization events that provide the ll- and dl-MeLan residues, respectively. The elimination domain of CoiSA likely dehydrates Thr57 to (E)-Dhb, which CoiC then converts to d-allo-l-MeLan. When CoiSA(ED) was coexpressed with CoiA1 and CoiC and a variant of CoiB that can still glutamylate but not eliminate, three dehydrations were still observed, but the cyclase was unable to form the three thioether macrocycles, presumably because the peptide now contained three (E)-Dhb residues. Thus, CoiC is only able to accept (E)-Dhb at position 57 and requires (Z)-Dhb at positions 43 and 50 for cyclization activity. The observation that CoiC is capable of cyclization of Cys51 onto (E)-Dhb57 but apparently cannot catalyze the addition of Cys47 and Cys55 to (E)-Dhb43 and 50, respectively, is suggestive of coevolution of CoiC with the appearance of CoiSA in the pathway. Conversely, when only CoiB was coexpressed with CoiA1 and CoiC, the dehydration process stalled at a single dehydration and one or two glutamylations. These findings suggest that either the syn-elimination of Thr57 by CoiSA(ED) is required for CoiB to complete its dehydration and/or that CoiC must first form the d-allo-l-MeLan for full CoiB activity. The data also suggest that after CoiSA/CoiC act on Thr57, glutamylation of Thr43 and Thr50 by the N-terminal domain of CoiB is followed by faster glutamate elimination by the GL domain of CoiB than by CoiSA(ED). These findings therefore suggest a highly choreographed set of biosynthetic reactions to make a complex ring pattern with high fidelity.
Orthologs of the CoiA1 peptide are much more common than orthologs of the OlvA peptide in the currently sequenced genomes (Figure 8B, Tables S7 and S8). Whether the coi-like BGCs evolved from the olv-like clusters by recruitment of a new GL domain or by gene duplication or whether the olv-like BGCs lost the gene for the syn-GL is a difficult question. Based on the preponderance of anti-GLs in diverse RiPP BGCs (Figure 7), it is likely that the ancestral enzyme catalyzed anti-elimination, but this hypothesis cannot be unambiguously verified at present. What is clear is that during evolution in Actinobacteria, two distinct BGCs with a common origin diverged and that stereochemistry was very likely a key determining factor. Determining the function of the coi and olv BGC products and how stereochemistry may alter the biological activities of these compounds is therefore of great interest. Investigations to answer these questions will first need to determine the cleavage site between the leader and core peptide, which is hampered by the absence of any reports detecting the products of these BGCs in their native producing organisms and the absence of a protease in the BGC that could provide insight regarding the start position of the final product.

Conclusions

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The coi BGC is highly similar in architecture to the previously investigated olv BGC with a key distinction being the presence of an additional GL domain that is fused to the MT CoiSA. We illustrate that mCoiA1 has a similar ring pattern to mOlvA with one less Lan ring and that it has different stereochemistry for two out of the three MeLan rings. The most pronounced difference between the two products is the recently discovered rare d-allo-l-MeLan diastereomer that is the result of the additional GL domain of CoiSA. This study illustrates an example of divergent evolution driven by stereochemistry, which in turn is likely to be correlated to the function of the final products.

Supporting Information

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The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acschembio.2c00492.

  • Glutamyl lyase SSN (ZIP)

  • Experimental procedures; extended experimental data and bioinformatic analysis; materials and methods; primers used in this study; observed and calculated m/z ratios for fragments of GluC-digested mCoiA1; NEM alkylation assay for CoiA1 co-expressed with CoiB-H994A, CoiC, and CoiSA(ED); LC-MS analysis of the GluC digestion product of mCoiA1 and variants; description table of orthologs obtained from PSI-BLAST of OlvA (PDF)

Accession Codes

NCBI protein accessions: CoiA1, WP_011031313.1; CoiB, WP_011031312.1; CoiC, WP_011031311.1; and CoiSA, WP_011031310.1.

Terms & Conditions

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Author Information

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  • Corresponding Author
    • Wilfred A. van der Donk - Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States;  Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States;  Orcidhttps://orcid.org/0000-0002-5467-7071;  Email: [email protected]
  • Author
    • Raymond Sarksian - Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, Illinois 61822, United States;  Orcidhttps://orcid.org/0000-0001-6940-6699
  • Funding

    This study was supported by the National Institutes of Health (Grant R37GM058822 to W.A.v.d.D.). The Bruker UltrafleXtreme MALDI TOF/TOF mass spectrometer was purchased in part with a grant from the National Center for Research Resources, National Institutes of Health (S10 RR027109 A). W.A.v.d.D. is an investigator of the Howard Hughes Medical Institute (HHMI).

  • Notes
    The authors declare no competing financial interest.

Acknowledgments

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The authors thank A. Ulanov at the Carver Biotechnology Center for help with GC-MS experiments and D. Sarlah (UIUC) for the use of a Kinetex Biphenyl column. They thank M. Simon for help with acquiring LC-MS data. This study is subject to HHMI’s Open Access to Publications policy. HHMI laboratory heads have previously granted a nonexclusive CC BY 4.0 license to the public and a sublicensable license to HHMI in their research articles. Pursuant to those licenses, the author-accepted manuscript of this article can be made freely available under a CC BY 4.0 license immediately upon publication.

References

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This article references 45 other publications.

  1. 1
    Harvey, A. L.; Edrada-Ebel, R.; Quinn, R. J. The re-emergence of natural products for drug discovery in the genomics era. Nature Rev. Drug Discovery 2015, 14, 111– 129,  DOI: 10.1038/nrd4510
    Google Scholar
    1
    The re-emergence of natural products for drug discovery in the genomics era
    Harvey, Alan L.; Edrada-Ebel, RuAngelie; Quinn, Ronald J.
    Nature Reviews Drug Discovery (2015), 14 (2), 111-129CODEN: NRDDAG; ISSN:1474-1776. (Nature Publishing Group)
    Natural products have been a rich source of compds. for drug discovery. However, their use has diminished in the past two decades, in part because of tech. barriers to screening natural products in high-throughput assays against mol. targets. Here, we review strategies for natural product screening that harness the recent tech. advances that have reduced these barriers. We also assess the use of genomic and metabolomic approaches to augment traditional methods of studying natural products, and highlight recent examples of natural products in antimicrobial drug discovery and as inhibitors of protein-protein interactions. The growing appreciation of functional assays and phenotypic screens may further contribute to a revival of interest in natural products for drug discovery.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhsVarsbg%253D&md5=b871148315dc0623683498fa15205b14
  2. 2
    Ortega, M. A.; van der Donk, W. A. New insights into the biosynthetic logic of ribosomally synthesized and post-translationally modified peptide natural products. Cell Chem. Biol. 2016, 23, 31– 44,  DOI: 10.1016/j.chembiol.2015.11.012
    Google Scholar
    2
    New Insights into the Biosynthetic Logic of Ribosomally Synthesized and Post-translationally Modified Peptide Natural Products
    Ortega, Manuel A.; van der Donk, Wilfred A.
    Cell Chemical Biology (2016), 23 (1), 31-44CODEN: CCBEBM; ISSN:2451-9448. (Cell Press)
    Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a large group of structurally diverse natural products. Their biol. activities and unique biosynthetic pathways have sparked a growing interest in RiPPs. Furthermore, the relatively low genetic complexity assocd. with RiPP biosynthesis makes them excellent candidates for synthetic biol. applications. This Review highlights recent developments in the understanding of the biosynthesis of several bacterial RiPP family members, the use of the RiPP biosynthetic machinery for generating novel macrocyclic peptides, and the implementation of tools designed to guide the discovery and characterization of novel RiPPs.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XntlSqsbo%253D&md5=1990f332ab2a2efb746aef7bb7718011
  3. 3
    Montalbán-López, M.; Scott, T. A.; Ramesh, S.; Rahman, I. R.; van Heel, A. J.; Viel, J. H.; Bandarian, V.; Dittmann, E.; Genilloud, O.; Goto, Y. New developments in RiPP discovery, enzymology and engineering. Nat. Prod. Rep. 2021, 38, 130– 239,  DOI: 10.1039/D0NP00027B
    Google Scholar
    3
    New developments in RiPP discovery, enzymology and engineering
    Montalban-Lopez, Manuel; Scott, Thomas A.; Ramesh, Sangeetha; Rahman, Imran R.; van Heel, Auke J.; Viel, Jakob H.; Bandarian, Vahe; Dittmann, Elke; Genilloud, Olga; Goto, Yuki; Grande Burgos, Maria Jose; Hill, Colin; Kim, Seokhee; Koehnke, Jesko; Latham, John A.; Link, A. James; Martinez, Beatriz; Nair, Satish K.; Nicolet, Yvain; Rebuffat, Sylvie; Sahl, Hans-Georg; Sareen, Dipti; Schmidt, Eric W.; Schmitt, Lutz; Severinov, Konstantin; Sussmuth, Roderich D.; Truman, Andrew W.; Wang, Huan; Weng, Jing-Ke; van Wezel, Gilles P.; Zhang, Qi; Zhong, Jin; Piel, Jorn; Mitchell, Douglas A.; Kuipers, Oscar P.; van der Donk, Wilfred A.
    Natural Product Reports (2021), 38 (1), 130-239CODEN: NPRRDF; ISSN:0265-0568. (Royal Society of Chemistry)
    A review. Up to June 2020 Ribosomally-synthesized and post-translationally modified peptides (RiPPs) are a large group of natural products. A community-driven review in 2013 described the emerging commonalities in the biosynthesis of RiPPs and the opportunities they offered for bioengineering and genome mining. Since then, the field has seen tremendous advances in understanding of the mechanisms by which nature assembles these compds., in engineering their biosynthetic machinery for a wide range of applications, and in the discovery of entirely new RiPP families using bioinformatic tools developed specifically for this compd. class. The First International Conference on RiPPs was held in 2019, and the meeting participants assembled the current review describing new developments since 2013. The review discusses the new classes of RiPPs that have been discovered, the advances in our understanding of the installation of both primary and secondary post-translational modifications, and the mechanisms by which the enzymes recognize the leader peptides in their substrates. In addn., genome mining tools used for RiPP discovery are discussed as well as various strategies for RiPP engineering. An outlook section presents directions for future research.
    >> More from SciFinder ®
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  4. 4
    Repka, L. M.; Chekan, J. R.; Nair, S. K.; van der Donk, W. A. Mechanistic understanding of lanthipeptide biosynthetic enzymes. Chem. Rev. 2017, 117, 5457– 5520,  DOI: 10.1021/acs.chemrev.6b00591
    Google Scholar
    4
    Mechanistic Understanding of Lanthipeptide Biosynthetic Enzymes
    Repka, Lindsay M.; Chekan, Jonathan R.; Nair, Satish K.; van der Donk, Wilfred A.
    Chemical Reviews (Washington, DC, United States) (2017), 117 (8), 5457-5520CODEN: CHREAY; ISSN:0009-2665. (American Chemical Society)
    A review. Lanthipeptides are ribosomally synthesized and post-translationally modified peptides (RiPPs) that display a wide variety of biol. activities, from antimicrobial to antiallodynic. Lanthipeptides that display antimicrobial activity are called lantibiotics. The post-translational modification reactions of lanthipeptides include dehydration of Ser and Thr residues to dehydroalanine and dehydrobutyrine, a transformation that is carried out in 3 unique ways in different classes of lanthipeptides. In a cyclization process, Cys residues then attack the dehydrated residues to generate the lanthionine and methyllanthionine thioether crosslinked amino acids from which lanthipeptides derive their name. The resulting polycyclic peptides have constrained conformations that confer their biol. activities. After installation of the characteristic thioether crosslinks, tailoring enzymes introduce addnl. post-translational modifications that are unique to each lanthipeptide and that fine-tune their activities and/or stability. This review focuses on studies published over the past decade that have provided much insight into the mechanisms of the enzymes that carry out the post-translational modifications.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhs1Oisr8%253D&md5=5bad9e545a5a1120c48a0b44c6ed6647
  5. 5
    Kodani, S.; Hudson, M. E.; Durrant, M. C.; Buttner, M. J.; Nodwell, J. R.; Willey, J. M. The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 11448– 11453,  DOI: 10.1073/pnas.0404220101
    Google Scholar
    5
    The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor
    Kodani, Shinya; Hudson, Michael E.; Durrant, Marcus C.; Buttner, Mark J.; Nodwell, Justin R.; Willey, Joanne M.
    Proceedings of the National Academy of Sciences of the United States of America (2004), 101 (31), 11448-11453CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
    SapB is a morphogenetic peptide that is important for aerial mycelium formation by the filamentous bacterium Streptomyces coelicolor. Prodn. of SapB commences during aerial mycelium formation and depends on most of the genes known to be required for the morphogenesis of aerial hyphae. Furthermore, the application of purified SapB to mutants blocked in morphogenesis restores their capacity to form aerial hyphae. Here, we present evidence that SapB is a lantibiotic-like peptide that is derived by posttranslational modification from the product of a gene (ramS) in the four-gene ram operon, which is under the control of the regulatory gene rams. We show that the product of another gene in the operon (ramC) contains a region that is similar to enzymes involved in the biosynthesis of lantibiotics, suggesting that it might be involved in the posttranslational processing of RamS. We conclude that SapB is derived from RamS through proteolytic cleavage and the introduction of four dehydroalanine residues and two lanthionine bridges. We provide an example of a morphogenetic role for an antibiotic-like mol.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXmvVKgurs%253D&md5=8da02864c5cbdc3f2e06067061f79603
  6. 6
    Kodani, S.; Lodato, M. A.; Durrant, M. C.; Picart, F.; Willey, J. M. SapT, a lanthionine-containing peptide involved in aerial hyphae formation in the Streptomycetes. Mol. Microbiol. 2005, 58, 1368– 1380,  DOI: 10.1111/j.1365-2958.2005.04921.x
    Google Scholar
    6
    SapT, a lanthionine-containing peptide involved in aerial hyphae formation in the streptomycetes
    Kodani, Shinya; Lodato, Michael A.; Durrant, Marcus C.; Picart, Francis; Willey, Joanne M.
    Molecular Microbiology (2005), 58 (5), 1368-1380CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Publishing Ltd.)
    The developmentally complex soil microbe Streptomyces tendae secretes a hydrophobic peptide that restored to developmental mutants of S. coelicolor the ability to raise aerial hyphae. The S. tendae peptide, SapT, has a lantibiotic structure and mol. modeling predicts that it is amphiphilic, making it structurally and functionally similar to the SapB peptide produced by S. coelicolor. However, SapT, which bears three β-Me lanthionine bridges and one lanthionine bridge and demonstrated limited antibiotic activity, is distinct from SapB. The amphiphilic nature of both SapT and SapB is required for their ability to serve as biosurfactants facilitating the emergence of newly formed aerial hyphae. Remarkably, SapB and SapT, and the fungal hydrophobin SC3 were shown to restore to a SapB-deficient S. coelicolor mutant the capacity to undergo complete morphogenesis, such that the extracellular addn. of protein resulted in sporulation. This suggests that the initiation of aerial growth may also indirectly trigger the signal transduction events needed for differentiation. These data imply that the prodn. of morphogenetic peptides may be common among the streptomycetes, but that while their ability to function as biosurfactants is conserved, their specific lantibiotic structure is not. Finally, the identification of a second lanthionine-contg. morphogenetic peptide suggests that lantibiotic structure and function may be more diverse than previously thought.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXhtlWmsL3K&md5=d5e31e9b605f7968ee90bf6b994638b4
  7. 7
    Willey, J. M.; Willems, A.; Kodani, S.; Nodwell, J. R. Morphogenetic surfactants and their role in the formation of aerial hyphae in Streptomyces coelicolor. Mol. Microbiol. 2006, 59, 731– 742,  DOI: 10.1111/j.1365-2958.2005.05018.x
    Google Scholar
    7
    Morphogenetic surfactants and their role in the formation of aerial hyphae in Streptomyces coelicolor
    Willey, Joanne M.; Willems, Andrew; Kodani, Shinya; Nodwell, Justin R.
    Molecular Microbiology (2006), 59 (3), 731-742CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Publishing Ltd.)
    A review. Withstanding environmental adversity and seeking optimal conditions for reprodn. are basic requirements for the survival of all organisms. Filamentous bacteria of the genus Streptomyces produce a remarkable cell type called the aerial hyphae that is central to its ability to meet both of these challenges. Recent advances have brought about a major shift in our understanding of the cell surface proteins that play important roles in the generation of these cells. Here we review our current understanding of one of these groups of proteins, the morphogenetic surfactants, with emphasis on the SapB protein of Streptomyces coelicolor.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28Xit1SgtLg%253D&md5=e362b3d2d36c1ba3d1b3aa0e02e560c9
  8. 8
    Férir, G.; Petrova, M. I.; Andrei, G.; Huskens, D.; Hoorelbeke, B.; Snoeck, R.; Vanderleyden, J.; Balzarini, J.; Bartoschek, S.; Brönstrup, M. The lantibiotic peptide labyrinthopeptin A1 demonstrates broad anti-HIV and anti-HSV activity with potential for microbicidal applications. PLoS One 2013, 8, e64010  DOI: 10.1371/journal.pone.0064010
    Google Scholar
    8
    The lantibiotic peptide labyrinthopeptin A1 demonstrates broad anti-HIV and anti-HSV activity with potential for microbicidal applications
    Ferir, Geoffrey; Petrova, Mariya I.; Andrei, Graciela; Huskens, Dana; Hoorelbeke, Bart; Snoeck, Robert; Vanderleyden, Jos; Balzarini, Jan; Bartoschek, Stefan; Broenstrup, Mark; Suessmuth, Roderich D.; Schols, Dominique
    PLoS One (2013), 8 (5), e64010CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)
    Lantibiotics are peptides, produced by bacteria, that contain the noncanonical amino acid lanthionine and many of them exhibit antibacterial activities. The labyrinthopeptin A1 (LabyA1) is a prototype peptide of a novel class of carbacyclic lantibiotics. Here, we extensively evaluated its broad-spectrum activity against HIV and HSV in vitro, studied its mechanism of action and evaluated potential microbicidal applications. LabyA1 exhibited a consistent and broad anti-HIV activity (EC50s: 0.70-3.3 μM) and anti-HSV activity (EC50s: 0.29-2.8 μM) in cell cultures. LabyA1 also inhibited viral cell-cell transmission between persistently HIV-infected T cells and uninfected CD4+ T cells (EC50: 2.5 μM) and inhibited the transmission of HIV captured by DC-SIGN+-cells to uninfected CD4+ T cells (EC50: 4.1 μM). Time-of-drug addn. studies revealed that LabyA1 acts as an entry inhibitor against HIV and HSV. Cellular and virus binding studies combined with SPR/FLIPR technol. showed that LabyA1 interacted with the HIV envelope protein gp120, but not with the HIV cellular receptors. LabyA1 also demonstrated additive to synergistic effects in its anti-HIV-1 and anti-HSV-2 activity with anti(retro)viral drugs in dual combinations such as tenofovir, acyclovir, saquinavir, raltegravir and enfuvirtide. LabyA1 can be considered as a novel lead peptide as it had profound antiviral activity against HIV and HSV. Pre-treatment of PBMCs with LabyA1 neither increased the expression of the activation markers CD69 and CD25, nor enhanced HIV replication, nor significantly induced various inflammatory cytokines/chemokines. LabyA1 also did not affect the growth of vaginal Lactobacilli populations. Based on the lack of toxicity on the vaginal Lactobacillus strains and its synergistic/additive profile in combination with clin. approved anti(retro)virals, it deserves further attention as a potential microbicide candidate in the prevention of sexual transmitted diseases.
    >> More from SciFinder ®
    https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXpsVKktLo%253D&md5=37d0f7354596b15f94c921c29bdf78b7
  9. 9
    Mohr, K. I.; Volz, C.; Jansen, R.; Wray, V.; Hoffmann, J.; Bernecker, S.; Wink, J.; Gerth, K.; Stadler, M.; Müller, R. Pinensins: the first antifungal lantibiotics. Angew. Chem., Int. Ed. 2015, 54, 11254– 11258,  DOI: 10.1002/anie.201500927
    Google Scholar
    9
    Pinensins: The first antifungal lantibiotics
    Mohr, Kathrin I.; Volz, Carsten; Jansen, Rolf; Wray, Victor; Hoffmann, Judith; Bernecker, Steffen; Wink, Joachim; Gerth, Klaus; Stadler, Marc; Mueller, Rolf
    Angewandte Chemie, International Edition (2015), 54 (38), 11254-11258CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)
    Lantibiotics (lanthionine-contg. antibiotics) from Gram-pos. bacteria typically exhibit activity against Gram-pos. bacteria. The activity and structure of pinensin A (1) and B (2), lantibiotics isolated from a native Gram-neg. producer Chitinophaga pinensis are described. Surprisingly, the pinensins were found to be highly active against many filamentous fungi and yeasts but show only weak antibacterial activity. To the best of our knowledge, lantibiotic fungicides have not been described before. An in-depth bioinformatic anal. of the biosynthetic gene cluster established the ribosomal origin of these compds. and identified candidate genes encoding all of the enzymes required for post-translational modification. Addnl. encoded functions enabled us to build up a hypothesis for the biosynthesis, export, sensing, and import of this intriguing lantibiotic.
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  10. 10
    Smith, T. E.; Pond, C. D.; Pierce, E.; Harmer, Z. P.; Kwan, J.; Zachariah, M. M.; Harper, M. K.; Wyche, T. P.; Matainaho, T. K.; Bugni, T. S. Accessing chemical diversity from the uncultivated symbionts of small marine animals. Nat. Chem. Biol. 2018, 14, 179– 185,  DOI: 10.1038/nchembio.2537
    Google Scholar
    10
    Accessing chemical diversity from the uncultivated symbionts of small marine animals
    Smith, Thomas E.; Pond, Christopher D.; Pierce, Elizabeth; Harmer, Zachary P.; Kwan, Jason; Zachariah, Malcolm M.; Harper, Mary Kay; Wyche, Thomas P.; Matainaho, Teatulohi K.; Bugni, Tim S.; Barrows, Louis R.; Ireland, Chris M.; Schmidt, Eric W.
    Nature Chemical Biology (2018), 14 (2), 179-185CODEN: NCBABT; ISSN:1552-4450. (Nature Research)
    Chem. drives many biol. interactions between the microbiota and host animals, yet it is often challenging to identify the chems. involved. This poses a problem, as such small mols. are excellent sources of potential pharmaceuticals, pretested by nature for animal compatibility. We discovered anti-HIV compds. from small, marine tunicates from the Eastern Fields of Papua New Guinea. Tunicates are a reservoir for new bioactive chems., yet their small size often impedes identification or even detection of the chems. within. We solved this problem by combining chem., metagenomics, and synthetic biol. to directly identify and synthesize the natural products. We show that these anti-HIV compds., the divamides, are a novel family of lanthipeptides produced by symbiotic bacteria living in the tunicate. Neighboring animal colonies contain structurally related divamides that differ starkly in their biol. properties, suggesting a role for biosynthetic plasticity in a native context wherein biol. interactions take place.
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  11. 11
    Garg, N.; Salazar-Ocampo, L. M. A.; van der Donk, W. A. In vitro activity of the nisin dehydratase NisB. Proc. Natl. Acad. Sci. U.S.A. 2013, 110, 7258– 7263,  DOI: 10.1073/pnas.1222488110
    Google Scholar
    11
    In vitro activity of the nisin dehydratase NisB
    Garg, Neha; Salazar-Ocampo, Luis M. A.; van der Donk, Wilfred A.
    Proceedings of the National Academy of Sciences of the United States of America (2013), 110 (18), 7258-7263, S7258/1-S7258/11CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
    The biosynthesis of several classes of ribosomally synthesized and posttranslationally modified peptides involves dehydration of serine and threonine residues. For class I lantibiotics, thiopeptides, and goadsporin, this dehydration is catalyzed by lanthionine biosynthetic enzyme B (LanB) or LanB-like proteins. Although LanB proteins have been studied since 1992, in vitro reconstitution of their dehydration activity has been elusive. We show here the in vitro activity of the dehydratase involved in the biosynthesis of the food preservative nisin (NisB). In vitro, NisB dehydrated its substrate peptide NisA eight times in the presence of glutamate, ATP, Mg2+, and the ribosomal/membrane fraction of bacterial cell ext. Mutation of 23 highly conserved residues of NisB identified a no. of amino acids that are essential for dehydration activity. In addn., these mutagenesis studies identified three mutants, R786A, R826A, and H961A, that result in multiple glutamylations of the NisA substrate. Glutamylation was obsd. during both Escherichia coli coexpression of NisA with these mutants and in vitro assays. Treatment of the glutamylated substrate with WT NisB results in dehydrated NisA, suggesting that the glutamylated peptide is an intermediate in dehydration. Collectively, these studies suggest that dehydration involves glutamylation of the side chains of Ser and Thr followed by elimination. The latter step has precedent in the virginiamycin resistance protein virginiamycin B lyase. These studies will facilitate investigation of other LanB proteins involved in the biosynthesis of lantibiotics, thiopeptides, and goadsporin.
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  12. 12
    Ortega, M. A.; Hao, Y.; Zhang, Q.; Walker, M. C.; van der Donk, W. A.; Nair, S. K. Structure and mechanism of the tRNA-dependent lantibiotic dehydratase NisB. Nature 2015, 517, 509– 512,  DOI: 10.1038/nature13888
    Google Scholar
    12
    Structure and mechanism of the tRNA-dependent lantibiotic dehydratase NisB
    Ortega, Manuel A.; Hao, Yue; Zhang, Qi; Walker, Mark C.; van der Donk, Wilfred A.; Nair, Satish K.
    Nature (London, United Kingdom) (2015), 517 (7535), 509-512CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)
    Lantibiotics are a class of peptide antibiotics that contain one or more thioether bonds. The lantibiotic, nisin, is an antimicrobial peptide that is widely used as a food preservative to combat food-borne pathogens. Nisin contains dehydroalanine and dehydrobutyrine residues that are formed by the dehydration of Ser/Thr by lantibiotic dehydratase NisB. Recent biochem. studies revealed that NisB glutamylates Ser/Thr side-chains as part of the dehydration process. However, the mol. mechanism by which NisB uses glutamate to catalyze dehydration remains unresolved. Here, the authors show that this process involves glutamyl-tRNAGlu to activate the Ser/Thr residues. In addn., the 2.9-Å crystal structure of Escherichia coli NisB in complex with its substrate peptide, nisin A, reveals the presence of 2 sep. domains that catalyze the Ser/Thr glutamylation and glutamate elimination steps. The co-crystal structure also provided insights into substrate recognition by lantibiotic dehydratases. These findings demonstrated an unexpected role for aminoacyl-tRNA in the formation of dehydroamino acids in lantibiotics, and served as a basis for the functional characterization of the many lantibiotic-like dehydratases involved in the biosynthesis of other classes of natural products.
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  13. 13
    Ortega, M. A.; Hao, Y.; Walker, M. C.; Donadio, S.; Sosio, M.; Nair, S. K.; van der Donk, W. A. Structure and tRNA specificity of MibB, a lantibiotic dehydratase from Actinobacteria involved in NAI-107 biosynthesis. Cell Chem. Biol. 2016, 23, 370– 380,  DOI: 10.1016/j.chembiol.2015.11.017
    Google Scholar
    13
    Structure and tRNA Specificity of MibB, a Lantibiotic Dehydratase from Actinobacteria Involved in NAI-107 Biosynthesis
    Ortega Manuel A; Hao Yue; Nair Satish K; van der Donk Wilfred A; Walker Mark C; van der Donk Wilfred A; Donadio Stefano; Sosio Margherita; Nair Satish K
    Cell chemical biology (2016), 23 (3), 370-380 ISSN:.
    Class I lantibiotic dehydratases dehydrate selected Ser/Thr residues of a precursor peptide. Recent studies demonstrated the requirement of glutamyl-tRNA(Glu) for Ser/Thr activation by one of these enzymes (NisB) from the Firmicute Lactococcus lactis. However, the generality of glutamyl-tRNA(Glu) usage and the tRNA specificity of lantibiotic dehydratases have not been established. Here we report the 2.7-ÅA resolution crystal structure, along with the glutamyl-tRNA(Glu) utilization of MibB, a lantibiotic dehydratase from the Actinobacterium Microbispora sp. 107891 involved in the biosynthesis of the clinical candidate NAI-107. Biochemical assays revealed nucleotides A73 and U72 within the tRNA(Glu) acceptor stem to be important for MibB glutamyl-tRNA(Glu) usage. Using this knowledge, an expression system for the production of NAI-107 analogs in Escherichia coli was developed, overcoming the inability of MibB to utilize E. coli tRNA(Glu). Our work provides evidence for a common tRNA(Glu)-dependent dehydration mechanism, paving the way for the characterization of lantibiotics from various phyla.
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  14. 14
    Bothwell, I. R.; Cogan, D. P.; Kim, T.; Reinhardt, C. J.; van der Donk, W. A.; Nair, S. K. Characterization of glutamyl-tRNA-dependent dehydratases using nonreactive substrate mimics. Proc. Natl. Acad. Sci. U.S.A. 2019, 116, 17245– 17250,  DOI: 10.1073/pnas.1905240116
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    14
    Characterization of glutamyl-tRNA-dependent dehydratases using nonreactive substrate mimics
    Bothwell, Ian R.; Cogan, Dillon P.; Kim, Terry; Reinhardt, Christopher J.; van der Donk, Wilfred A.; Nair, Satish K.
    Proceedings of the National Academy of Sciences of the United States of America (2019), 116 (35), 17245-17250CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
    The peptide natural product nisin has been used as a food preservative for 6 decades with minimal development of resistance. Nisin contains the unusual amino acids dehydroalanine and dehydrobutyrine, which are posttranslationally installed by class I lanthipeptide dehydratases (LanBs) on a linear peptide substrate through an unusual glutamyl-tRNA-dependent dehydration of Ser and Thr. To date, little is known about how LanBs catalyze the transfer of glutamate from charged tRNAGlu to the peptide substrate, or how they carry out the subsequent elimination of the peptide-glutamyl adducts to afford dehydro amino acids. Here, we describe the synthesis of inert analogs that mimic substrate glutamyl-tRNAGlu and the glutamylated peptide intermediate, and det. the crystal structures of 2 LanBs in complex with each of these compds. Mutational studies were used to characterize the function of the glutamylation and glutamate elimination active-site residues identified through the structural anal. These combined studies provide insights into the mechanisms of substrate recognition, glutamylation, and glutamate elimination by LanBs to effect a net dehydration reaction of Ser and Thr.
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  15. 15
    Li, B.; Yu, J. P. J.; Brunzelle, J. S.; Moll, G. N.; van der Donk, W. A.; Nair, S. K. Structure and mechanism of the lantibiotic cyclase involved in nisin biosynthesis. Science 2006, 311, 1464– 1467,  DOI: 10.1126/science.1121422
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    15
    Structure and Mechanism of the Lantibiotic Cyclase Involved in Nisin Biosynthesis
    Li, Bo; Yu, John Paul J.; Brunzelle, Joseph S.; Moll, Gert N.; van der Donk, Wilfred A.; Nair, Satish K.
    Science (Washington, DC, United States) (2006), 311 (5766), 1464-1467CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)
    Nisin is a posttranslationally modified antimicrobial peptide that is widely used as a food preservative. It contains five cyclic thioethers of varying sizes that are installed by a single enzyme, NisC. Reported here are the in vitro reconstitution of the cyclization process and the x-ray crystal structure of the NisC enzyme. The structure reveals similarities in fold and substrate activation with mammalian farnesyl transferases, suggesting that human homologs of NisC posttranslationally modify a cysteine of a protein substrate.
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  16. 16
    Li, B.; van der Donk, W. A. Identification of essential catalytic residues of the cyclase NisC involved in the biosynthesis of nisin. J. Biol. Chem. 2007, 282, 21169– 21175,  DOI: 10.1074/jbc.M701802200
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    16
    Identification of Essential Catalytic Residues of the Cyclase NisC Involved in the Biosynthesis of Nisin
    Li, Bo; van der Donk, Wilfred A.
    Journal of Biological Chemistry (2007), 282 (29), 21169-21175CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)
    Nisin is a post-translationally modified antimicrobial peptide that has been widely used in the food industry for several decades. It contains five cyclic thioether cross-links of varying sizes that are installed by a single enzyme, NisC, that catalyzes the addn. of cysteines to dehydroamino acids. The recent X-ray crystal structure of NisC has provided the first insights into the catalytic residues responsible for the cyclization step during nisin biosynthesis. In this study, the conserved residues His212, Arg280, Asp141, and Tyr285 as well as the ligands to the zinc in the active site (Cys284, Cys330, and His331) were substituted by site-directed mutagenesis. Binding studies showed that all mutants had similar affinities for NisA. Activity assays showed that whereas His212 and Asp141 were essential for correct cyclization as judged by the antimicrobial activity of the final product, Arg280 and Tyr285 were not. Mutation of zinc ligands to alanine also abolished the enzymic activity, and these mutant proteins were shown to contain decreased levels of zinc. These results show that the zinc is essential for activity and support a model in which the zinc is used to activate the cysteines in the substrate for nucleophilic attack. These findings also argue against an essential role of Arg280 and Tyr285 as an active site general acid/base in the mechanism of cyclization.
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  17. 17
    Mukherjee, S.; Huo, L.; Thibodeaux, G. N.; van der Donk, W. A. Synthesis and bioactivity of diastereomers of the virulence lanthipeptide cytolysin. Org. Lett. 2016, 18, 6188– 6191,  DOI: 10.1021/acs.orglett.6b03246
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    17
    Synthesis and bioactivity of diastereomers of the virulence lanthipeptide cytolysin
    Mukherjee, Subha; Huo, Liujie; Thibodeaux, Gabrielle N.; van der Donk, Wilfred A.
    Organic Letters (2016), 18 (23), 6188-6191CODEN: ORLEF7; ISSN:1523-7052. (American Chemical Society)
    Cytolysin, a two-component lanthipeptide comprising cytolysin S (CylLS'') and cytolysin L (CylLL''), is the only family member to exhibit lytic activity against mammalian cells in addn. to synergistic antimicrobial activity. A subset of the thioether cross-links of CylLS'' and CylLL'' have LL stereochem. instead of the canonical DL stereochem. in all previously characterized lanthipeptides. The synthesis of a CylLS'' variant with DL stereochem. is reported. Its antimicrobial activity was found to be decreased, but not its lytic activity against red blood cells. Hence, the unusual LL stereochem. is not responsible for the lytic activity.
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  18. 18
    Knerr, P. J.; van der Donk, W. A. Chemical synthesis of the lantibiotic lacticin 481 reveals the importance of lanthionine stereochemistry. J. Am. Chem. Soc. 2013, 135, 7094– 7097,  DOI: 10.1021/ja4014024
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    18
    Chemical synthesis of the lantibiotic Lacticin 481 reveals the importance of lanthionine stereochemistry
    Knerr, Patrick J.; van der Donk, Wilfred A.
    Journal of the American Chemical Society (2013), 135 (19), 7094-7097CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lantibiotics are a family of antibacterial peptide natural products characterized by the post-translational installation of the thioether-contg. amino acids lanthionine and methyllanthionine. Until recently, only a single naturally occurring stereochem. configuration for each of these cross-links was known. The discovery of lantibiotics with alternative lanthionine and methyllanthionine stereochem. has prompted an investigation of its importance to biol. activity. Here, solid-supported chem. synthesis enabled the total synthesis of the lantibiotic lacticin 481 and analogs contg. cross-links with non-native stereochem. configurations. Biol. evaluation revealed that these alterations abolished the antibacterial activity in all of the analogs, revealing the crit. importance of the enzymically installed stereochem. for the biol. activity of lacticin 481.
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  19. 19
    Tang, W.; van der Donk, W. A. The sequence of the enterococcal cytolysin imparts unusual lanthionine stereochemistry. Nat. Chem. Biol. 2013, 9, 157– 159,  DOI: 10.1038/nchembio.1162
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    19
    The sequence of the enterococcal cytolysin imparts unusual lanthionine stereochemistry
    Tang, Weixin; van der Donk, Wilfred A.
    Nature Chemical Biology (2013), 9 (3), 157-159CODEN: NCBABT; ISSN:1552-4450. (Nature Publishing Group)
    The enterococcal cytolysin is a two-component lantibiotic of unknown structure with hemolytic activity that is important for virulence. We prepd. cytolysin by coexpression of each precursor peptide with the synthetase CylM in Escherichia coli and characterized its structure. Unexpectedly, cytolysin is to our knowledge the first example of a lantibiotic contg. lanthionine and methyllanthionine structures with different stereochemistries in the same peptide. The stereochem. is detd. by the sequence of the substrate peptide.
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  20. 20
    Tang, W.; Jiménez-Osés, G.; Houk, K. N.; van der Donk, W. A. Substrate control in stereoselective lanthionine biosynthesis. Nat. Chem. 2015, 7, 57– 64,  DOI: 10.1038/nchem.2113
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    20
    Substrate control in stereoselective lanthionine biosynthesis
    Tang, Weixin; Jimenez-Oses, Gonzalo; Houk, K. N.; van der Donk, Wilfred A.
    Nature Chemistry (2015), 7 (1), 57-64CODEN: NCAHBB; ISSN:1755-4330. (Nature Publishing Group)
    Enzymes are typically highly stereoselective catalysts that enforce a reactive conformation on their native substrates. We report here a rare example in which the substrate controls the stereoselectivity of an enzyme-catalyzed Michael-type addn. during the biosynthesis of lanthipeptides. These natural products contain thioether crosslinks formed by a cysteine attack on dehydrated Ser and Thr residues. We demonstrate that several lanthionine synthetases catalyze highly selective anti-addns. in which the substrate (and not the enzyme) dets. whether the addn. occurs from the re or si face. A single point mutation in the peptide substrate completely inverted the stereochem. outcome of the enzymic modification. Quantum mech. calcns. reproduced the exptl. obsd. selectivity and suggest that conformational restraints imposed by the amino-acid sequence on the transition states det. the face selectivity of the Michael-type cyclization.
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  21. 21
    Tang, W.; Thibodeaux, G. N.; van der Donk, W. A. The enterococcal cytolysin synthetase coevolves with substrate for stereoselective lanthionine synthesis. ACS Chem. Biol. 2016, 11, 2438– 2446,  DOI: 10.1021/acschembio.6b00397
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    21
    The enterococcal cytolysin synthetase coevolves with substrate for stereoselective lanthionine synthesis
    Tang, Weixin; Thibodeaux, Gabrielle N.; van der Donk, Wilfred A.
    ACS Chemical Biology (2016), 11 (9), 2438-2446CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)
    Stereochem. control is crit. in natural product biosynthesis. For ribosomally synthesized and post-translationally modified peptides (RiPPs), the mechanism(s) by which stereoselectivity is achieved is still poorly understood. In this work, we focused on the stereoselective lanthionine synthesis in lanthipeptides, a major class of RiPPs formed by the addn. of Cys residues to dehydroalanine (Dha) or dehydrobutyrine (Dhb). Nonenzymic cyclization of the small subunit of a virulence lanthipeptide, the enterococcal cytolysin, resulted in the native modified peptide as the major product, suggesting that both regioselectivity and stereoselectivity are inherent to the dehydrated peptide sequence. These results support previous computational studies that a Dhx-Dhx-Xxx-Xxx-Cys motif (Dhx = Dha or Dhb; Xxx = any amino acid except Dha, Dhb, and Cys) preferentially cyclizes by attack on the Re face of Dha or Dhb. Characterization of the stereochem. of the products formed enzymically with substrate mutants revealed that the lanthionine synthetase actively reinforces Re face attack. These findings support the hypothesis of substrate-controlled selectivity in lanthionine synthesis but also reveal likely coevolution of substrates and lanthionine synthetases to ensure the stereoselective synthesis of lanthipeptides with defined biol. activities.
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  22. 22
    Garg, N.; Goto, Y.; Chen, T.; van der Donk, W. A. Characterization of the stereochemical configuration of lanthionines formed by the lanthipeptide synthetase GeoM. Biopolymers 2016, 106, 834– 842,  DOI: 10.1002/bip.22876
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    22
    Characterization of the stereochemical configuration of lanthionines formed by the lanthipeptide synthetase GeoM
    Garg, Neha; Goto, Yuki; Chen, Ting; van der Donk, Wilfred A.
    Biopolymers (2016), 106 (6), 834-842CODEN: BIPMAA; ISSN:0006-3525. (John Wiley & Sons, Inc.)
    The biosynthesis of the class II lanthipeptide geobacillin II was reconstituted in vitro. The purified precursor peptide was modified by the lanthipeptide synthetase GeoM at temps. ranging between 37 and 80°C demonstrating the thermostability of the enzyme. Geobacillin II shares with cytolysin, haloduracin, and carnolysin a DhxDhxXxxXxxCys motif (Dhx = dehydroalanine or dehydrobutyrine) as precursor to its N-terminal A-ring. Like in these other three lantibiotics, the lanthionine in the A-ring of geobacillin II had the LL stereochem. configuration as shown by chiral gas chromatog./mass spectrometry. Various analogs of geobacillin II were produced using co-expression of mutants of the precursor peptide GeoAII and the synthetase GeoM in Escherichia coli. The findings in this study suggest that the stereochem. outcome of the A-ring in geobacillin II is not solely dependent on the peptide sequence as previously suggested for haloduracin © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 834-842, 2016.
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  23. 23
    Lohans, C. T.; Li, J. L.; Vederas, J. C. Structure and biosynthesis of carnolysin, a homologue of enterococcal cytolysin with D-amino acids. J. Am. Chem. Soc. 2014, 136, 13150– 13153,  DOI: 10.1021/ja5070813
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    23
    Structure and biosynthesis of carnolysin, a homolog of enterococcal cytolysin with D-amino acids
    Lohans, Christopher T.; Li, Jessica L.; Vederas, John C.
    Journal of the American Chemical Society (2014), 136 (38), 13150-13153CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lantibiotics are a group of highly post-translationally modified bacterial antimicrobial peptides characterized by the presence of the thioether-contg. amino acids lanthionine and methyllanthionine. Carnobacterium maltaromaticum C2 was found to produce a two-component lantibiotic homologous to enterococcal cytolysin. Through tandem mass spectrometry and NMR spectroscopy, the post-translational modifications of carnolysin were established, and the topologies of the lanthionine and methyllanthionine rings were detd. Chiral GC-MS anal. revealed that, like cytolysin, carnolysin contained lanthionine and methyllanthionine residues of unusual stereochem. Carnolysin, unlike cytolysin, was shown to contain D-alanine and unprecedented D-aminobutyrate derived from serine and threonine, resp. Carnolysin was heterologously expressed in Escherichia coli, demonstrating that reductase CrnJ is involved in the formation of the D-amino acids.
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  24. 24
    Sarksian, R.; Hegemann, J. D.; Simon, M. A.; Acedo, J. Z.; van der Donk, W. A. Unexpected methyllanthionine stereochemistry in the morphogenetic lanthipeptide SapT. J. Am. Chem. Soc. 2022, 144, 6373– 6382,  DOI: 10.1021/jacs.2c00517
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    24
    Unexpected methyllanthionine stereochemistry in the morphogenetic lanthipeptide SapT
    Sarksian, Raymond; Hegemann, Julian D.; Simon, Max A.; Acedo, Jeella Z.; van der Donk, Wilfred A.
    Journal of the American Chemical Society (2022), 144 (14), 6373-6382CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lanthipeptides are polycyclic peptides characterized by the presence of lanthionine (Lan) and/or methyllanthionine (MeLan). They are members of the ribosomally synthesized and post-translationally modified peptides (RiPPs). The stereochem. configuration of (Me)Lan crosslinks is important for the bioactivity of lanthipeptides. To date, MeLan residues in characterized lanthipeptides have either the 2S,3S or 2R,3R stereochem. Herein, we reconstituted in Escherichia coli the biosynthetic pathway toward SapT, a class I lanthipeptide that exhibits morphogenetic activity. Through the synthesis of stds., the heterologously produced peptide was shown to possess three MeLan residues with the 2S,3R stereochem. (D-allo-L-MeLan), the first time such stereochem. has been obsd. in a lanthipeptide. Bioinformatic anal. of the biosynthetic enzymes suggests this stereochem. may also be present in other lanthipeptides. Anal. of another gene cluster in Streptomyces coelicolor that is widespread in actinobacteria confirmed another example of D-allo-L-MeLan and verified the bioinformatic prediction. We propose a mechanism for the origin of the unexpected stereochem. and provide support using site-directed mutagenesis.
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  25. 25
    Bothwell, I. R.; Caetano, T.; Sarksian, R.; Mendo, S.; van der Donk, W. A. Structural analysis of class I lanthipeptides from Pedobacter lusitanus NL19 reveals an unusual ring pattern. ACS Chem. Biol. 2021, 16, 1019– 1029,  DOI: 10.1021/acschembio.1c00106
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    25
    Structural Analysis of Class I Lanthipeptides from Pedobacter lusitanus NL19 Reveals an Unusual Ring Pattern
    Bothwell, Ian R.; Caetano, Tania; Sarksian, Raymond; Mendo, Sonia; van der Donk, Wilfred A.
    ACS Chemical Biology (2021), 16 (6), 1019-1029CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)
    Lanthipeptides are ribosomally synthesized and post-translationally modified peptide natural products characterized by the presence of lanthionine and methyllanthionine crosslinked amino acids formed by dehydration of Ser/Thr residues followed by conjugate addn. of Cys to the resulting dehydroamino acids. Class I lanthipeptide dehydratases use glutamyl-tRNAGlu as a cosubstrate to glutamylate Ser/Thr followed by glutamate elimination. A vast majority of lanthipeptides identified from class I synthase systems were from Gram-pos. bacteria. Herein, the authors report the heterologous expression and modification in Escherichia coli of two lanthipeptides from the Gram-neg. Bacteroidetes Pedobacter lusitanus NL19. These peptides are representative of a group of compds. frequently encoded in Pedobacter genomes. Structural characterization of the lanthipeptides revealed a novel ring pattern as well as an unusual LL-lanthionine stereochem. configuration and a cyclase that lacks the canonical zinc ligands found in most LanC enzymes.
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  26. 26
    Acedo, J. Z.; Bothwell, I. R.; An, L.; Trouth, A.; Frazier, C.; van der Donk, W. A. O-methyltransferase-mediated incorporation of a β-amino acid in lanthipeptides. J. Am. Chem. Soc. 2019, 141, 16790– 16801,  DOI: 10.1021/jacs.9b07396
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    26
    O-Methyltransferase-Mediated Incorporation of a β-Amino Acid in Lanthipeptides
    Acedo, Jeella Z.; Bothwell, Ian R.; An, Linna; Trouth, Abby; Frazier, Clara; van der Donk, Wilfred A.
    Journal of the American Chemical Society (2019), 141 (42), 16790-16801CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lanthipeptides represent a large class of cyclic natural products defined by the presence of lanthionine (Lan) and methyllanthionine (MeLan) cross-links. With the advances in DNA sequencing technologies and genome mining tools, new biosynthetic enzymes capable of installing unusual structural features are continuously being discovered. In this study, we investigated an O-methyltransferase that is a member of the most prominent auxiliary enzyme family assocd. with class I lanthipeptide biosynthetic gene clusters. Despite the prevalence of these enzymes, their function has not been established. Herein, we demonstrate that the O-methyltransferase OlvSA encoded in the olv gene cluster from Streptomyces olivaceus NRRL B-3009 catalyzes the rearrangement of a highly conserved aspartate residue to a β-amino acid, isoaspartate, in the lanthipeptide OlvA(BCSA). We elucidated the NMR soln. structure of the GluC-digested peptide, OlvA(BCSA)GluC, which revealed a unique ring topol. comprising four interlocking rings and positions the isoaspartate residue in a solvent exposed loop that is stabilized by a MeLan ring. Gas chromatog.-mass spectrometry anal. further indicated that OlvA(BCSA) contains two DL-MeLan rings and two Lan rings with an unusual LL-stereochem. Lastly, in vitro reconstitution of OlvSA activity showed that it is a leader peptide-independent and S-adenosyl methionine-dependent O-methyltransferase that mediates the conversion of a highly conserved aspartate residue in a cyclic substrate into a succinimide, which is hydrolyzed to generate an Asp or isoAsp contg. peptide. This overall transformation converts an α-amino acid into a β-amino acid in a ribosomally synthesized peptide, via an electrophilic intermediate that may be the intended product.
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  27. 27
    Marsh, A. J.; O’Sullivan, O.; Ross, R. P.; Cotter, P. D.; Hill, C. In silico analysis highlights the frequency and diversity of type 1 lantibiotic gene clusters in genome sequenced bacteria. BMC Genomics 2010, 11, 679  DOI: 10.1186/1471-2164-11-679
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    In silico analysis highlights the frequency and diversity of type 1 lantibiotic gene clusters in genome sequenced bacteria
    Marsh, Alan J.; O'Sullivan, Orla; Ross, R. Paul; Cotter, Paul D.; Hill, Colin
    BMC Genomics (2010), 11 (), 679CODEN: BGMEET; ISSN:1471-2164. (BioMed Central Ltd.)
    Background: Lantibiotics are lanthionine-contg., post-translationally modified antimicrobial peptides. These peptides have significant, but largely untapped, potential as preservatives and chemotherapeutic agents. Type 1 lantibiotics are those in which lanthionine residues are introduced into the structural peptide (LanA) through the activity of sep. lanthionine dehydratase (LanB) and lanthionine synthetase (LanC) enzymes. Here we take advantage of the conserved nature of LanC enzymes to devise an in silico approach to identify potential lantibiotic-encoding gene clusters in genome sequenced bacteria. Results: In total 49 novel type 1 lantibiotic clusters were identified which unexpectedly were assocd. with species, genera and even phyla of bacteria which have not previously been assocd. with lantibiotic prodn. Conclusions: Multiple type 1 lantibiotic gene clusters were identified at a frequency that suggests that these antimicrobials are much more widespread than previously thought. These clusters represent a rich repository which can yield a large no. of valuable novel antimicrobials and biosynthetic enzymes.
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  28. 28
    Walker, M. C.; Eslami, S. M.; Hetrick, K. J.; Ackenhusen, S. E.; Mitchell, D. A.; van der Donk, W. A. Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family. BMC Genomics 2020, 21, 387  DOI: 10.1186/s12864-020-06785-7
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    28
    Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family
    Walker, Mark C.; Eslami, Sara M.; Hetrick, Kenton J.; Ackenhusen, Sarah E.; Mitchell, Douglas A.; van der Donk, Wilfred A.
    BMC Genomics (2020), 21 (1), 387CODEN: BGMEET; ISSN:1471-2164. (BioMed Central Ltd.)
    Lanthipeptides belong to the ribosomally synthesized and post-translationally modified peptide group of natural products and have a variety of biol. activities ranging from antibiotics to antinociceptives. These peptides are cyclized through thioether crosslinks and can bear other secondary post-translational modifications. While lanthipeptide biosynthetic gene clusters can be identified by the presence of genes encoding characteristic enzymes involved in the post-translational modification process, locating the precursor peptides encoded within these clusters is challenging due to their short length and high sequence variability, which limits the high-throughput exploration of lanthipeptide biosynthesis. To address this challenge, we enhanced the predictive capabilities of Rapid ORF Description & Evaluation Online (RODEO) to identify members of all four known classes of lanthipeptides. Using RODEO, we mined over 100,000 bacterial and archaeal genomes in the RefSeq database. We identified nearly 8500 lanthipeptide precursor peptides. These precursor peptides were identified in a broad range of bacterial phyla as well as the Euryarchaeota phylum of archaea. Bacteroidetes were found to encode a large no. of these biosynthetic gene clusters, despite making up a relatively small portion of the genomes in this dataset. A no. of these precursor peptides are similar to those of previously characterized lanthipeptides, but even more were not, including potential antibiotics. One such new antimicrobial lanthipeptide was purified and characterized. Addnl., examn. of the biosynthetic gene clusters revealed that enzymes installing secondary post-translational modifications are more widespread than initially thought. Lanthipeptide biosynthetic gene clusters are more widely distributed and the precursor peptides encoded within these clusters are more diverse than previously appreciated, demonstrating that the lanthipeptide sequence-function space remains largely underexplored.
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  29. 29
    Bentley, S. D.; Chater, K. F.; Cerdeno-Tarraga, A. M.; Challis, G. L.; Thomson, N. R.; James, K. D.; Harris, D. E.; Quail, M. A.; Kieser, H.; Harper, D. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature 2002, 417, 141– 147,  DOI: 10.1038/417141a
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    Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2)
    Bentley, S. D.; Chater, K. F.; Cerdeno-Tarraga, A.-M.; Challis, G. L.; Thomson, N. R.; James, K. D.; Harris, D. E.; Quail, M. A.; Kieser, H.; Harper, D.; Bateman, A.; Brown, S.; Chandra, G.; Chen, C. W.; Collins, M.; Cronin, A.; Fraser, A.; Goble, A.; Hidalgo, J.; Hornsby, T.; Howarth, S.; Huang, C.-H.; Kieser, T.; Larke, L.; Murphy, L.; Oliver, K.; O'Neil, S.; Rabbinowitsch, E.; Rajandream, M.-A.; Rutherford, K.; Rutter, S.; Seeger, K.; Saunders, D.; Sharp, S.; Squares, R.; Squares, S.; Taylor, K.; Warren, T.; Wietzorrek, A.; Woodward, J.; Barrell, B. G.; Parkhill, J.; Hopwood, D. A.
    Nature (London, United Kingdom) (2002), 417 (6885), 141-147CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)
    Streptomyces coelicolor is a representative of the group of soil-dwelling, filamentous bacteria responsible for producing most natural antibiotics used in human and veterinary medicine. The 8,667,507 base pair linear chromosome of this organism, contg. the largest no. of genes so far discovered in a bacterium, is reported. The 7825 predicted genes include more than 20 clusters coding for known or predicted secondary metabolites. The genome contains an unprecedented proportion of regulatory genes, predominantly those likely to be involved in responses to external stimuli and stresses, and many duplicated gene sets that may represent 'tissue-specific' isoforms operating in different phases of colonial development, a unique situation for a bacterium. An ancient synteny was revealed between the central 'core' of the chromosome and the whole chromosome of pathogens Mycobacterium tuberculosis and Corynebacterium diphtheriae. The genome sequence will greatly increase our understanding of microbial life in the soil as well as aiding the generation of new drug candidates by genetic engineering. The complete sequence is deposited in GenBank/EMBL under accession no. AL645882. [This abstr. record is one of two records for this document necessitated by the large no. of index entries required to fully index the document and publication system constraints.].
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  30. 30
    Challis, G. L. Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways. J. Ind. Microbiol. Biotechnol. 2014, 41, 219– 232,  DOI: 10.1007/s10295-013-1383-2
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    Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways
    Challis, Gregory L.
    Journal of Industrial Microbiology & Biotechnology (2014), 41 (2), 219-232CODEN: JIMBFL; ISSN:1367-5435. (Springer)
    A review. Streptomyces, and related genera of Actinobacteria, are renowned for their ability to produce antibiotics and other bioactive natural products with a wide range of applications in medicine and agriculture. Streptomyces coelicolor A3(2) is a model organism that has been used for more than five decades to study the genetic and biochem. basis for the prodn. of bioactive metabolites. In 2002, the complete genome sequence of S. coelicolor was published. This greatly accelerated progress in understanding the biosynthesis of metabolites known or suspected to be produced by S. coelicolor and revealed that streptomycetes have far greater potential to produce bioactive natural products than suggested by classical bioassay-guided isolation studies. In this article, efforts to exploit the S. coelicolor genome sequence for the discovery of novel natural products and biosynthetic pathways are summarized.
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  31. 31
    Ozaki, T.; Nishiyama, M.; Kuzuyama, T. Novel tryptophan metabolism by a potential gene cluster that is widely distributed among actinomycetes. J. Biol. Chem. 2013, 288, 9946– 9956,  DOI: 10.1074/jbc.M112.436451
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    31
    Novel tryptophan metabolism by a potential gene cluster that is widely distributed among Actinomycetes
    Ozaki, Taro; Nishiyama, Makoto; Kuzuyama, Tomohisa
    Journal of Biological Chemistry (2013), 288 (14), 9946-9956CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)
    The characterization of potential gene clusters is a promising strategy for the identification of novel natural products and the expansion of structural diversity. However, there are often difficulties in identifying potential metabolites because their biosynthetic genes are either silenced or expressed only at a low level. Here, we report the identification of a novel metabolite that is synthesized by a potential gene cluster contg. an indole prenyltransferase gene (SCO7467) and a flavin-dependent monooxygenase (FMO) gene (SCO7468), which were mined from the genome of Streptomyces coelicolor A3(2). We introduced these two genes into the closely related Streptomyces lividans TK23 and analyzed the culture broths of the transformants. This process allowed us to identify a novel metabolite, 5-dimethylallylindole-3-acetonitrile (5-DMAIAN) that was overproduced in the transformant. Biochem. characterization of the recombinant SCO7467 and SCO7468 demonstrated the novel l-tryptophan metab. leading to 5-DMAIAN. SCO7467 catalyzes the prenylation of l-tryptophan to form 5-dimethylallyl-l-tryptophan (5-DMAT). This enzyme is the first actinomycetes prenyltransferase known to catalyze the addn. of a dimethylallyl group to the C-5 of tryptophan. SCO7468 then catalyzes the conversion of 5-DMAT into 5-dimethylallylindole-3-acetaldoxime (5-DMAIAOx). An aldoxime-forming reaction catalyzed by the FMO enzyme was also identified for the first time in this study. Finally, dehydration of 5-DMAIAOx presumably occurs to yield 5-DMAIAN. This study provides insight into the biosynthesis of prenylated indoles that have been purified from actinomycetes.
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    Nodwell, J. R. Microbe profile: Streptomyces coelicolor: a burlesque of pigments and phenotypes. Microbiology 2019, 165, 953– 955,  DOI: 10.1099/mic.0.000821
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    Microbe profile: Streptomyces coelicolor: a burlesque of pigments and phenotypes
    Nodwell, Justin R.
    Microbiology (London, United Kingdom) (2019), 165 (9), 953-955CODEN: MROBEO; ISSN:1465-2080. (Microbiology Society)
    The streptomycetes are soil-dwelling bacteria that are found in soil everywhere on Earth: the mol. geosmin, which they produce as part of their life cycle, is what gives soil its familiar 'earthy' smell. The species is best known for the prodn. of biol. active small mols. called 'natural products'. These mols. are the source of most of our antibiotics and anti-fungals, as well as many other drugs. The streptomycetes have a filamentous form rather than the more familiar rod-shaped spirochete and coccoid forms. They exhibit a complex life cycle and sporulation mechanism involving several differentiated cell types, each having specific roles in the colony life history. Streptomyces coelicolor is an important model system for this genus - research on this bacterium has provided foundational information for all of these fascinating processes.
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    Lautru, S.; Deeth, R. J.; Bailey, L. M.; Challis, G. L. Discovery of a new peptide natural product by Streptomyces coelicolor genome mining. Nat. Chem. Biol. 2005, 1, 265– 269,  DOI: 10.1038/nchembio731
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    33
    Discovery of a new peptide natural product by Streptomyces coelicolor genome mining
    Lautru, Sylvie; Deeth, Robert J.; Bailey, Lianne M.; Challis, Gregory L.
    Nature Chemical Biology (2005), 1 (5), 265-269CODEN: NCBABT; ISSN:1552-4450. (Nature Publishing Group)
    Analyses of microbial genome sequences reveal numerous examples of gene clusters encoding proteins typically involved in complex natural product biosynthesis but not assocd. with the prodn. of known natural products. In Streptomyces coelicolor M145 there are several gene clusters encoding new nonribosomal peptide synthetase (NRPS) systems not assocd. with known metabolites. Application of structure-based models for substrate recognition by NRPS adenylation domains predicts the amino acids incorporated into the putative peptide products of these systems, but the accuracy of these predictions is untested. Here we report the isolation and structure detn. of the new tris-hydroxamate tetrapeptide iron chelator coelichelin from S. coelicolor using a genome mining approach guided by substrate predictions for the trimodular NRPS CchH, and we show that this enzyme, which lacks a C-terminal thioesterase domain, together with a homolog of enterobactin esterase (CchJ), are required for coelichelin biosynthesis. These results demonstrate that accurate prediction of adenylation domain substrate selectivity is possible and raise intriguing mechanistic questions regarding the assembly of a tetrapeptide by a trimodular NRPS.
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  34. 34
    Song, L.; Barona-Gomez, F.; Corre, C.; Xiang, L.; Udwary, D. W.; Austin, M. B.; Noel, J. P.; Moore, B. S.; Challis, G. L. Type III polyketide synthase beta-ketoacyl-ACP starter unit and ethylmalonyl-CoA extender unit selectivity discovered by Streptomyces coelicolor genome mining. J. Am. Chem. Soc. 2006, 128, 14754– 14755,  DOI: 10.1021/ja065247w
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    Type III Polyketide Synthase β-Ketoacyl-ACP Starter Unit and Ethylmalonyl-CoA Extender Unit Selectivity Discovered by Streptomyces coelicolor Genome Mining
    Song, Lijiang; Barona-Gomez, Francisco; Corre, Christophe; Xiang, Longkuan; Udwary, Daniel W.; Austin, Michael B.; Noel, Joseph P.; Moore, Bradley S.; Challis, Gregory L.
    Journal of the American Chemical Society (2006), 128 (46), 14754-14755CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Polyketide synthases (PKSs) are involved in the biosynthesis of many important natural products. In bacteria, type III PKSs typically catalyze iterative decarboxylation and condensation reactions of malonyl-CoA building blocks in the biosynthesis of polyhydroxyarom. products. Here it is shown that Gcs, a type III PKS encoded by the sco7221 ORF of the bacterium Streptomyces coelicolor, is required for biosynthesis of the germicidin family of 3,6-dialkyl-4-hydroxypyran-2-one natural products. Evidence consistent with Gcs-catalyzed elongation of specific β-ketoacyl-ACP products of the fatty acid synthase FabH with ethyl- or methylmalonyl-CoA in the biosynthesis of germicidins is presented. Selectivity for β-ketoacyl-ACP starter units and ethylmalonyl-CoA as an extender unit is unprecedented for type III PKSs, suggesting these enzymes may be capable of utilizing a far wider range of starter and extender units for natural product assembly than believed until now.
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    Lin, X.; Hopson, R.; Cane, D. E. Genome mining in Streptomyces coelicolor: molecular cloning and characterization of a new sesquiterpene synthase. J. Am. Chem. Soc. 2006, 128, 6022– 6023,  DOI: 10.1021/ja061292s
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    Genome Mining in Streptomyces coelicolor: Molecular Cloning and Characterization of a New Sesquiterpene Synthase
    Lin, Xin; Hopson, Russell; Cane, David E.
    Journal of the American Chemical Society (2006), 128 (18), 6022-6023CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    The terpene synthase encoded by the SCO5222 (SC7E4.19) gene of Streptomyces coelicolor was cloned by PCR and expressed in Escherichia coli as an N-terminal-His6-tag protein. Incubation of the recombinant protein, SCO5222p, with farnesyl diphosphate (1, FPP) in the presence of Mg(II) gave a new sesquiterpene, (+)-epi-isozizaene (2), whose structure and stereochem. were detd. by a combination of 1H, 13C, COSY, HMQC, HMBC, and NOESY NMR. The steady-state kinetic parameters were kcat 0.049±0.001 s-1 and a Km (FPP) of 147±14 nM. Individual incubations of recombinant epi-isozizaene synthase with [1,1-2H2]FPP (1a), (1R)-[1-2H]-FPP (1b), and (1S)-[1-2H]-FPP (1c) and NMR anal. of the resulting deuterated epi-isozizaenes supported an isomerization-cyclization-rearrangement mechanism involving the intermediacy of (3R)-nerolidyl diphosphate (3).
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  36. 36
    Corre, C.; Song, L.; O’Rourke, S.; Chater, K. F.; Challis, G. L. 2-Alkyl-4-hydroxymethylfuran-3-carboxylic acids, antibiotic production inducers discovered by Streptomyces coelicolor genome mining. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 17510– 17515,  DOI: 10.1073/pnas.0805530105
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    2-Alkyl-4-hydroxymethylfuran-3-carboxylic acids, antibiotic production inducers discovered by Streptomyces coelicolor genome mining
    Corre, Christophe; Song, Lijiang; O'Rourke, Sean; Chater, Keith F.; Challis, Gregory L.
    Proceedings of the National Academy of Sciences of the United States of America (2008), 105 (45), 17510-17515CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
    All of the genetic elements necessary for the prodn. of the antibiotic methylenomycin (Mm) and its regulation are contained within the 22-kb mmy-mmf gene cluster, which is located on the 356-kb linear plasmid SCP1 of Streptomyces coelicolor A3(2). A putative operon of 3 genes within this gene cluster, mmfLHP, was proposed to direct the biosynthesis of an A-factor-like signaling mol., which could play a role in the regulation of Mm biosynthesis. The mmfLHP operon was expressed under the control of its native promoter in S. coelicolor M512, a host lacking the SCP1 plasmid, and the ability to produce prodiginine and actinorhodin antibiotics. Comparative metabolic profiling led to the identification and structure elucidation of a family of 5 new 2-alkyl-4-hydroxymethylfuran-3-carboxylic acids (AHFCAs), collectively termed Mm furans (MMFs), as the products of the mmfLHP genes. MMFs specifically induce the prodn. of the Mm antibiotics in S. coelicolor. Comparative genomics analyses and searches of the natural product chem. literature indicated that other streptomycetes may produce AHFCAs, suggesting that they could form a general class of antibiotic biosynthesis inducers in Streptomyces species, with analogous functions to the better known γ-butyrolactone regulatory mols.
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  37. 37
    Wu, C.; Ichinose, K.; Choi, Y. H.; van Wezel, G. P. Aromatic polyketide GTRI-02 is a previously unidentified product of the act gene cluster in Streptomyces coelicolor A3(2). ChemBioChem 2017, 18, 1428– 1434,  DOI: 10.1002/cbic.201700107
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    Aromatic Polyketide GTRI-02 is a Previously Unidentified Product of the act Gene Cluster in Streptomyces coelicolor A3(2)
    Wu, Changsheng; Ichinose, Koji; Choi, Young Hae; van Wezel, Gilles P.
    ChemBioChem (2017), 18 (14), 1428-1434CODEN: CBCHFX; ISSN:1439-4227. (Wiley-VCH Verlag GmbH & Co. KGaA)
    The biosynthesis of arom. polyketides derived from type II polyketide synthases (PKSs) is complex, and it is not uncommon that highly similar gene clusters give rise to diverse structural architectures. The act biosynthetic gene cluster (BGC) of the model actinomycete Streptomyces coelicolor A3(2) is an archetypal type II PKS. Here we show that the act BGC also specifies the arom. polyketide GTRI-02 (1) and propose a mechanism for the biogenesis of its 3,4-dihydronaphthalen-1(2H)-one backbone. Polyketide 1 was also produced by Streptomyces sp. MBT76 after activation of the act-like qin gene cluster by overexpression of the pathway-specific activator. Mining of this strain also identified dehydroxy-GTRI-02 (2), which most likely originated from dehydration of 1 during the isolation process. This work shows that even extensively studied model gene clusters such as act of S. coelicolor can still produce new chem., offering new perspectives for drug discovery.
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  38. 38
    Li, B.; Cooper, L. E.; van der Donk, W. A. In Vitro Studies of Lantibiotic Biosynthesis. In Methods Enzymol.; Elsevier, 2009; Vol. 458, pp 533– 558.
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    There is no corresponding record for this reference.
  39. 39
    Küsters, E.; Allgaier, H.; Jung, G.; Bayer, E. Resolution of sulphur-containing amino acids by chiral phase gas chromatography. Chromatographia 1984, 18, 287– 293,  DOI: 10.1007/BF02259079
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    39
    Resolution of sulfur-containing amino acids by chiral phase gas chromatography
    Kuesters, E.; Allgaier, H.; Jung, G.; Bayer, E.
    Chromatographia (1984), 18 (6), 287-93CODEN: CHRGB7; ISSN:0009-5893.
    Me esters of the pentafluoropropionyl derivs. of lanthionines, cystathionines, and β-methyllanthionines were resolved by gas chromatog. on glass capillaries coated with the chiral stationary phase N-propionyl-L-valine tert-butylamide-polysiloxane (Chirasil-Val) within 35 min. L-Cystathionine elutes before its D-enantiomer in contrast to the usual order of emergence on an L-phase. The method was applied to the polypeptide antibiotic nisin, which contains meso-lanthionine and (2S,3S,6R)-3-methyllanthionine. N-Pentafluoropropionyl-S-alkylthiocysteine Me esters (alkyl = Me, Et, n- and iso-Pr, n- and sec-Bu, n-octyl, neo-pentyl, cyclohexyl, benzyl, tolyl) were sepd. on Chirasil-Val within 30 min. The identity of all derivs. was shown by combined gas chromatog.-mass spectrometry.
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  40. 40
    Liu, W.; Chan, A. S. H.; Liu, H.; Cochrane, S. A.; Vederas, J. C. Solid supported chemical syntheses of both components of the lantibiotic lacticin 3147. J. Am. Chem. Soc. 2011, 133, 14216– 14219,  DOI: 10.1021/ja206017p
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    40
    Solid supported chemical syntheses of both components of the lantibiotic lacticin 3147
    Liu, Wei; Chan, Alice S. H.; Liu, Hongqiang; Cochrane, Stephen A.; Vederas, John C.
    Journal of the American Chemical Society (2011), 133 (36), 14216-14219CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lantibiotics are antimicrobial peptides produced by bacteria. Some are employed for food preservation, whereas others have therapeutic potential due to their activity against organisms resistant to current antibiotics. They are ribosomally synthesized and posttranslationally modified by dehydration of serine and threonine residues followed by attack of thiols of cysteines to form monosulfide lanthionine and methyllanthionine rings, resp. Chem. synthesis of peptide analogs is a powerful method to verify stereochem. and access structure-activity relationships. However, solid supported synthesis of lantibiotics has been difficult due to problems in generating lanthionines and methyllanthionines with orthogonal protection and good stereochem. control. We report the solid-phase syntheses of both peptides of a two-component lantibiotic, lacticin 3147. Both successive and interlocking ring systems were synthesized on-resin, thereby providing a general methodol. for this family of natural products.
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  41. 41
    Zallot, R.; Oberg, N.; Gerlt, J. A. The EFI web resource for genomic enzymology tools: Leveraging protein, genome, and metagenome databases to discover novel enzymes and metabolic pathways. Biochemistry 2019, 58, 4169– 4182,  DOI: 10.1021/acs.biochem.9b00735
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    41
    The EFI Web Resource for Genomic Enzymology Tools: Leveraging Protein, Genome, and Metagenome Databases to Discover Novel Enzymes and Metabolic Pathways
    Zallot, Remi; Oberg, Nils; Gerlt, John A.
    Biochemistry (2019), 58 (41), 4169-4182CODEN: BICHAW; ISSN:0006-2960. (American Chemical Society)
    The assignment of functions to uncharacterized proteins discovered in genome projects requires easily accessible tools and computational resources for large-scale, user-friendly leveraging of the protein, genome, and metagenome databases by experimentalists. This article describes the web resource developed by the Enzyme Function Initiative (EFI; accessed at https://efi.igb.illinois.edu/) that provides "genomic enzymol." tools ("web tools") for (1) generating sequence similarity networks (SSNs) for protein families (EFI-EST); (2) analyzing and visualizing genome context of the proteins in clusters in SSNs (in genome neighborhood networks, GNNs, and genome neighborhood diagrams, GNDs) (EFI-GNT); and (3) prioritizing uncharacterized SSN clusters for functional assignment based on metagenome abundance (chem. guided functional profiling, CGFP) (EFI-CGFP). The SSNs generated by EFI-EST are used as the input for EFI-GNT and EFI-CGFP, enabling easy transfer of information among the tools. The networks are visualized and analyzed using Cytoscape, a widely used desktop application; GNDs and CGFP heatmaps summarizing metagenome abundance are viewed within the tools. We provide a detailed example of the integrated use of the tools with an anal. of glycyl radical enzyme superfamily (IPR004184) found in the human gut microbiome. This anal. demonstrates that (1) SwissProt annotations are not always correct, (2) large-scale genome context analyses allow the prediction of novel metabolic pathways, and (3) metagenome abundance can be used to identify/prioritize uncharacterized proteins for functional investigation.
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  42. 42
    Gerlt, J. A.; Bouvier, J. T.; Davidson, D. B.; Imker, H. J.; Sadkhin, B.; Slater, D. R.; Whalen, K. L. Enzyme Function Initiative-Enzyme Similarity Tool (EFI-EST): A web tool for generating protein sequence similarity networks. Biochim. Biophys. Acta, Proteins Proteomics 2015, 1854, 1019– 1037,  DOI: 10.1016/j.bbapap.2015.04.015
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    Enzyme function initiative-enzyme similarity tool (EFI-EST): A web tool for generating protein sequence similarity networks
    Gerlt, John A.; Bouvier, Jason T.; Davidson, Daniel B.; Imker, Heidi J.; Sadkhin, Boris; Slater, David R.; Whalen, Katie L.
    Biochimica et Biophysica Acta, Proteins and Proteomics (2015), 1854 (8), 1019-1037CODEN: BBAPBW; ISSN:1570-9639. (Elsevier B. V.)
    A review. The Enzyme Function Initiative, an NIH/NIGMS-supported Large-Scale Collaborative Project (EFI; U54GM093342; http://enzymefunction.org/), is focused on devising and disseminating bioinformatics and computational tools as well as exptl. strategies for the prediction and assignment of functions (in vitro activities and in vivo physiol./metabolic roles) to uncharacterized enzymes discovered in genome projects. Protein sequence similarity networks (SSNs) are visually powerful tools for analyzing sequence relationships in protein families (H.J. Atkinson, J.H. Morris, T.E. Ferrin, and P.C. Babbitt, PLoS One 2009, 4, e4345). However, the members of the biol./biomedical community have not had access to the capability to generate SSNs for their "favorite" protein families. In this article we announce the EFI-EST (Enzyme Function Initiative-Enzyme Similarity Tool) web tool (http://efi.igb.illinois.edu/efi-est/) that is available without cost for the automated generation of SSNs by the community. The tool can create SSNs for the "closest neighbors" of a user-supplied protein sequence from the UniProt database (Option A) or of members of any user-supplied Pfam and/or InterPro family (Option B). We provide an introduction to SSNs, a description of EFI-EST, and a demonstration of the use of EFI-EST to explore sequence-function space in the OMP decarboxylase superfamily (PF00215). This article is designed as a tutorial that will allow members of the community to use the EFI-EST web tool for exploring sequence/function space in protein families.
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  43. 43
    Le, T.; Fouque, K. J. D.; Santos-Fernandez, M.; Navo, C. D.; Jiménez-Osés, G.; Sarksian, R.; Fernandez-Lima, F. A.; van Der Donk, W. A. Substrate sequence controls regioselectivity of lanthionine formation by ProcM. J. Am. Chem. Soc. 2021, 143, 18733– 18743,  DOI: 10.1021/jacs.1c09370
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    Substrate Sequence Controls Regioselectivity of Lanthionine Formation by ProcM
    Le, Tung; Jeanne Dit Fouque, Kevin; Santos-Fernandez, Miguel; Navo, Claudio D.; Jimenez-Oses, Gonzalo; Sarksian, Raymond; Fernandez-Lima, Francisco Alberto; van der Donk, Wilfred A.
    Journal of the American Chemical Society (2021), 143 (44), 18733-18743CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
    Lanthipeptides belong to the family of ribosomally synthesized and post-translationally modified peptides (RiPPs). The (methyl)lanthionine crosslinks characteristic to lanthipeptides are essential for their stability and bioactivities. In most bacteria, lanthipeptides are maturated from single precursor peptides encoded in the corresponding biosynthetic gene clusters. However, cyanobacteria engage in combinatorial biosynthesis and encode as many as 80 substrate peptides with highly diverse sequences that are modified by a single lanthionine synthetase into lanthipeptides of different lengths and ring patterns. It is puzzling how a single enzyme could exert control over the cyclization processes of such a wide range of substrates. Here, the authors used a library of ProcA3.3 precursor peptide variants and show that it is not the enzyme ProcM but rather its substrate sequences that det. the regioselectivity of lanthionine formation. The authors also demonstrate the utility of trapped ion mobility spectrometry-tandem mass spectrometry (TIMS-MS/MS) as a fast and convenient method to efficiently sep. lanthipeptide constitutional isomers, particularly in cases where the isomers cannot be resolved by conventional liq. chromatog. The authors' data allowed identification of factors that are important for the cyclization outcome, but also showed that there are no easily identifiable predictive rules for all sequences. The authors' findings provide a platform for future deep learning approaches to allow such prediction of ring patterns of products of combinatorial biosynthesis.
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  44. 44
    Bobeica, S. C.; Zhu, L.; Acedo, J. Z.; Tang, W.; van der Donk, W. A. Structural determinants of macrocyclization in substrate-controlled lanthipeptide biosynthetic pathways. Chem. Sci. 2020, 11, 12854– 12870,  DOI: 10.1039/D0SC01651A
    Google Scholar
    44
    Structural determinants of macrocyclization in substrate-controlled lanthipeptide biosynthetic pathways
    Bobeica, Silvia C.; Zhu, Lingyang; Acedo, Jeella Z.; Tang, Weixin; van der Donk, Wilfred A.
    Chemical Science (2020), 11 (47), 12854-12870CODEN: CSHCCN; ISSN:2041-6520. (Royal Society of Chemistry)
    Lanthipeptides are characterized by thioether crosslinks formed by post-translational modifications. The cyclization process that favors a single ring pattern over many other possible ring patterns has been the topic of much speculation. Recent studies suggest that for some systems the cyclization pattern and stereochem. is detd. not by the enzyme, but by the sequence of the precursor peptide. However, the factors that govern the outcome of the cyclization process are not understood. This study presents the three-dimensional structures of seven lanthipeptides detd. by NMR spectroscopy, including five prochlorosins and the two peptides that make up cytolysin, a virulence factor produced by Enterococcus faecalis that is directly linked to human disease. These peptides were chosen because their substrate sequence dets. either the ring pattern (prochlorosins) or the stereochem. of cyclization (cytolysins). We present the structures of prochlorosins 1.1, 2.1, 2.8, 2.10 and 2.11, the first three-dimensional structures of prochlorosins. Our findings provide insights into the mol. determinants of cyclization as well as why some prochlorosins may be better starting points for library generation than others. The structures of the large and small subunits of the enterococcal cytolysin show that these peptides have long helical stretches, a rare observation for lanthipeptides characterized to date. These helixes may explain their pore forming activity and suggest that the small subunit may recognize a mol. target followed by recruitment of the large subunit to span the membrane.
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  45. 45
    Altschul, S. F.; Madden, T. L.; Schaffer, A. A.; Zhang, J.; Zhang, Z.; Miller, W.; Lipman, D. J. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997, 25, 3389– 3402,  DOI: 10.1093/nar/25.17.3389
    Google Scholar
    45
    Gapped BLAST and PSI-BLAST: a new generation of protein database search programs
    Altschul, Stephen F.; Madden, Thomas L.; Schaffer, Alejandro A.; Zhang, Jinghui; Zhang, Zheng; Miller, Webb; Lipman, David J.
    Nucleic Acids Research (1997), 25 (17), 3389-3402CODEN: NARHAD; ISSN:0305-1048. (Oxford University Press)
    The BLAST programs are widely used tools for searching protein and DNA databases for sequence similarities. For protein comparisons, a variety of definitional, algorithmic and statistical refinements described here permits the execution time of the BLAST programs to be decreased substantially while enhancing their sensitivity to weak similarities. A new criterion for triggering the extension of word hits, combined with a new heuristic for generating gapped alignments, yields a gapped BLAST program that runs at approx. three times the speed of the original. In addn., a method is introduced for automatically combining statistically significant alignments produced by BLAST into a position-specific score matrix, and searching the database using this matrix. The resulting Position-Specific Iterated BLAST (PSI-BLAST) program runs at approx. the same speed per iteration as gapped BLAST, but in many cases is much more sensitive to weak but biol. relevant sequence similarities. PSI-BLAST is used to uncover several new and interesting members of the BRCT superfamily. The source code for the new BLAST programs is available by anonymous ftp from the machine ncbi.nlm.nih.gov, within the directory 'blast', and the programs may be run from NCBIs web site at http://www.ncbi.nlm.nih.gov/.
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  1. Emily K. Desormeaux, Garrett J. Barksdale, Wilfred A. van der Donk. Kinetic Analysis of Cyclization by the Substrate-Tolerant Lanthipeptide Synthetase ProcM. ACS Catalysis 2024, 14 (24) , 18310-18321. https://doi.org/10.1021/acscatal.4c06216
  2. Ross Rayne Vermeulen, Anton Du Preez van Staden, Tracey Ollewagen, Leonardo Joaquim van Zyl, Youran Luo, Wilfred A. van der Donk, Leon Milner Theodore Dicks, Carine Smith, Marla Trindade. Initial Characterization of the Viridisins’ Biological Properties. ACS Omega 2024, 9 (29) , 31832-31841. https://doi.org/10.1021/acsomega.4c03149
  3. Youran Luo, Shuyun Xu, Autumn M. Frerk, Wilfred A. van der Donk. Facile Method for Determining Lanthipeptide Stereochemistry. Analytical Chemistry 2024, 96 (4) , 1767-1773. https://doi.org/10.1021/acs.analchem.3c04958
  4. Guannan Zhong. Cytochromes P450 Associated with the Biosyntheses of Ribosomally Synthesized and Post-translationally Modified Peptides. ACS Bio & Med Chem Au 2023, 3 (5) , 371-388. https://doi.org/10.1021/acsbiomedchemau.3c00026
  5. Li Cao, Truc Do, Angela Zhu, Jianshu Duan, Nathan Alam, A. James Link. Genome Mining and Discovery of Imiditides, a Family of RiPPs with a Class-Defining Aspartimide Modification. Journal of the American Chemical Society 2023, 145 (34) , 18834-18845. https://doi.org/10.1021/jacs.3c03991
  6. Emily K. Desormeaux, Wilfred A. van der Donk. Kinetic Analysis of Lanthipeptide Cyclization by Substrate-Tolerant ProcM. 2024https://doi.org/10.1101/2024.05.16.594612
  7. Youran Luo, Shuyun Xu, Wilfred A. van der Donk. A Facile Method for Determining Lanthipeptide Stereochemistry. 2023https://doi.org/10.1101/2023.10.26.564210
  8. Christopher J. Thibodeaux. The conformationally dynamic structural biology of lanthipeptide biosynthesis. Current Opinion in Structural Biology 2023, 81 , 102644. https://doi.org/10.1016/j.sbi.2023.102644
  9. Li Cao, Truc Do, Angela D. Zhu, Nathan Alam, A. James Link. Genome Mining and Discovery of Imiditides, a Novel Family of RiPPs with a Class-defining Aspartimide Modification. 2023https://doi.org/10.1101/2023.04.07.536058
  10. Hongshuang Liu, Senbiao Fang, Lin Zhao, Xiao Men, Haibo Zhang. A Single Active-Site Mutagenesis Confers Enhanced Activity and/or Changed Product Distribution to a Pentalenene Synthase from Streptomyces sp. PSKA01. Bioengineering 2023, 10 (3) , 392. https://doi.org/10.3390/bioengineering10030392
  11. Raymond Sarksian, Lingyang Zhu, Wilfred A. van der Donk. syn -Elimination of glutamylated threonine in lanthipeptide biosynthesis. Chemical Communications 2023, 59 (9) , 1165-1168. https://doi.org/10.1039/D2CC06345J
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  • Abstract

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    Figure 1

    Figure 1. (A) Maturation of lanthipeptides proceeds through dehydration of Ser/Thr residues to generate the corresponding dehydroamino acids dehydroalanine and dehydrobutyrine (Dha, Dhb). Cyclization of Cys side chains onto Dha/Dhb yields (Me)Lan. (B) Four possible MeLan diastereomers can be generated as shown.

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    Figure 2

    Figure 2. (A) Comparison of coi and olv class I lanthipeptide BGCs. The coi BGC encodes an additional GL and protein of unknown function when compared to the olv BGC. (B) Sequence alignment of the precursor peptides encoded in both BGCs. Residues that are known to be modified in OlvA are colored. (C) Sequence alignment of GL domains illustrates that the additional GL domain in CoiSA is similar to SptBb.

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    Figure 3

    Figure 3. Liquid chromatography-MS (LC-MS) analysis of the GluC digestion product of mCoiA1 (mCoiA1GluC). (A) High-resolution electrospray ionization-MS (ESI-MS) analysis. (B) Tandem ESI-MS analysis. Fragmentation results are consistent with an N-terminal MeLan and two C-terminal overlapping MeLan.

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    Figure 4

    Figure 4. Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS analysis of coexpression experiments in E. coli. Coexpressed proteins are listed on each panel. Asterisks indicate deamination products that are commonly generated in MALDI-TOF mass spectra at these masses.

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    Figure 5

    Figure 5. Gas chromatography-MS (GC-MS) analysis of derivatized MeLan residues (m/z = 379) obtained from mCoiA1 mutant peptides using a chiral stationary phase. Coinjections with stereochemically pure derivatized dl- and ll-MeLan confirm the presence of dl-MeLan in mCoiA1-T43S and ll-MeLan in mCoiA1-T50S. (A) Derivatized MeLan from WT mCoiA1 (top), synthetic standards, and coinjections of the products of mCoiA1 with the synthetic standards. (B) Derivatized MeLan from mCoiA1-T43S (top), synthetic standards, and coinjections of the products of mCoiA1-T43S with the synthetic standards. (C) Derivatized MeLan from mCoiA1-T50S (top), synthetic standards, and coinjections of the products of mCoiA1-T50S with the synthetic standards. (D) Structures of derivatized dl- and ll-MeLan standards.

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    Figure 6

    Figure 6. MALDI-TOF MS analysis of CoiA1 coexpression with CoiB, CoiC, and CoiSA(ED) WT or CoiSA(ED) mutants. Asterisks indicate deamination products that are commonly generated in MALDI-TOF mass spectra at these masses.

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    Figure 7

    Figure 7. SSN analysis of GLs. All colored groups indicate GLs within class I lanthipeptide BGCs. Full-length LanB dehydratases that catalyze anti-elimination are shown in black, and anti-GLs within split LanB systems in blue. Syn-GLs that are part of a split LanB system that are fused to a methyltransferase or that are present as stand-alone proteins are depicted in purple. GLs within BGCs of other nonlanthipeptide or hybrid RiPPs are in gray. A select number of characterized GLs are labeled including the thiopeptide GLs TbtC and LazF. For brief summaries of the biosynthetic genes in each group, see Table S6. The cytoscape file for the SSN is provided as the Supporting Information.

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    Figure 8

    Figure 8. (A) Comparison of the structures of GluC-digested mCoiA1 and mOlvA to highlight differences and similarities in ring patterns and stereochemistry. Abu, 2-aminobutyric acid. The N-terminal Lan ring in mOlvA is absent in mCoiA1, although the corresponding Ser residue (gold) is present. (B) Sequence LOGO made using orthologs of the core peptide region of OlvA and CoiA1 identified by BLAST analysis (45) to highlight conserved residues (OlvA total sequences = 43, CoiA1 total sequences = 301).

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  • References

    This article references 45 other publications.

    1. 1
      Harvey, A. L.; Edrada-Ebel, R.; Quinn, R. J. The re-emergence of natural products for drug discovery in the genomics era. Nature Rev. Drug Discovery 2015, 14, 111– 129,  DOI: 10.1038/nrd4510
      1
      The re-emergence of natural products for drug discovery in the genomics era
      Harvey, Alan L.; Edrada-Ebel, RuAngelie; Quinn, Ronald J.
      Nature Reviews Drug Discovery (2015), 14 (2), 111-129CODEN: NRDDAG; ISSN:1474-1776. (Nature Publishing Group)
      Natural products have been a rich source of compds. for drug discovery. However, their use has diminished in the past two decades, in part because of tech. barriers to screening natural products in high-throughput assays against mol. targets. Here, we review strategies for natural product screening that harness the recent tech. advances that have reduced these barriers. We also assess the use of genomic and metabolomic approaches to augment traditional methods of studying natural products, and highlight recent examples of natural products in antimicrobial drug discovery and as inhibitors of protein-protein interactions. The growing appreciation of functional assays and phenotypic screens may further contribute to a revival of interest in natural products for drug discovery.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhsVarsbg%253D&md5=b871148315dc0623683498fa15205b14
    2. 2
      Ortega, M. A.; van der Donk, W. A. New insights into the biosynthetic logic of ribosomally synthesized and post-translationally modified peptide natural products. Cell Chem. Biol. 2016, 23, 31– 44,  DOI: 10.1016/j.chembiol.2015.11.012
      2
      New Insights into the Biosynthetic Logic of Ribosomally Synthesized and Post-translationally Modified Peptide Natural Products
      Ortega, Manuel A.; van der Donk, Wilfred A.
      Cell Chemical Biology (2016), 23 (1), 31-44CODEN: CCBEBM; ISSN:2451-9448. (Cell Press)
      Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a large group of structurally diverse natural products. Their biol. activities and unique biosynthetic pathways have sparked a growing interest in RiPPs. Furthermore, the relatively low genetic complexity assocd. with RiPP biosynthesis makes them excellent candidates for synthetic biol. applications. This Review highlights recent developments in the understanding of the biosynthesis of several bacterial RiPP family members, the use of the RiPP biosynthetic machinery for generating novel macrocyclic peptides, and the implementation of tools designed to guide the discovery and characterization of novel RiPPs.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XntlSqsbo%253D&md5=1990f332ab2a2efb746aef7bb7718011
    3. 3
      Montalbán-López, M.; Scott, T. A.; Ramesh, S.; Rahman, I. R.; van Heel, A. J.; Viel, J. H.; Bandarian, V.; Dittmann, E.; Genilloud, O.; Goto, Y. New developments in RiPP discovery, enzymology and engineering. Nat. Prod. Rep. 2021, 38, 130– 239,  DOI: 10.1039/D0NP00027B
      3
      New developments in RiPP discovery, enzymology and engineering
      Montalban-Lopez, Manuel; Scott, Thomas A.; Ramesh, Sangeetha; Rahman, Imran R.; van Heel, Auke J.; Viel, Jakob H.; Bandarian, Vahe; Dittmann, Elke; Genilloud, Olga; Goto, Yuki; Grande Burgos, Maria Jose; Hill, Colin; Kim, Seokhee; Koehnke, Jesko; Latham, John A.; Link, A. James; Martinez, Beatriz; Nair, Satish K.; Nicolet, Yvain; Rebuffat, Sylvie; Sahl, Hans-Georg; Sareen, Dipti; Schmidt, Eric W.; Schmitt, Lutz; Severinov, Konstantin; Sussmuth, Roderich D.; Truman, Andrew W.; Wang, Huan; Weng, Jing-Ke; van Wezel, Gilles P.; Zhang, Qi; Zhong, Jin; Piel, Jorn; Mitchell, Douglas A.; Kuipers, Oscar P.; van der Donk, Wilfred A.
      Natural Product Reports (2021), 38 (1), 130-239CODEN: NPRRDF; ISSN:0265-0568. (Royal Society of Chemistry)
      A review. Up to June 2020 Ribosomally-synthesized and post-translationally modified peptides (RiPPs) are a large group of natural products. A community-driven review in 2013 described the emerging commonalities in the biosynthesis of RiPPs and the opportunities they offered for bioengineering and genome mining. Since then, the field has seen tremendous advances in understanding of the mechanisms by which nature assembles these compds., in engineering their biosynthetic machinery for a wide range of applications, and in the discovery of entirely new RiPP families using bioinformatic tools developed specifically for this compd. class. The First International Conference on RiPPs was held in 2019, and the meeting participants assembled the current review describing new developments since 2013. The review discusses the new classes of RiPPs that have been discovered, the advances in our understanding of the installation of both primary and secondary post-translational modifications, and the mechanisms by which the enzymes recognize the leader peptides in their substrates. In addn., genome mining tools used for RiPP discovery are discussed as well as various strategies for RiPP engineering. An outlook section presents directions for future research.
      >> More from SciFinder ®
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    4. 4
      Repka, L. M.; Chekan, J. R.; Nair, S. K.; van der Donk, W. A. Mechanistic understanding of lanthipeptide biosynthetic enzymes. Chem. Rev. 2017, 117, 5457– 5520,  DOI: 10.1021/acs.chemrev.6b00591
      4
      Mechanistic Understanding of Lanthipeptide Biosynthetic Enzymes
      Repka, Lindsay M.; Chekan, Jonathan R.; Nair, Satish K.; van der Donk, Wilfred A.
      Chemical Reviews (Washington, DC, United States) (2017), 117 (8), 5457-5520CODEN: CHREAY; ISSN:0009-2665. (American Chemical Society)
      A review. Lanthipeptides are ribosomally synthesized and post-translationally modified peptides (RiPPs) that display a wide variety of biol. activities, from antimicrobial to antiallodynic. Lanthipeptides that display antimicrobial activity are called lantibiotics. The post-translational modification reactions of lanthipeptides include dehydration of Ser and Thr residues to dehydroalanine and dehydrobutyrine, a transformation that is carried out in 3 unique ways in different classes of lanthipeptides. In a cyclization process, Cys residues then attack the dehydrated residues to generate the lanthionine and methyllanthionine thioether crosslinked amino acids from which lanthipeptides derive their name. The resulting polycyclic peptides have constrained conformations that confer their biol. activities. After installation of the characteristic thioether crosslinks, tailoring enzymes introduce addnl. post-translational modifications that are unique to each lanthipeptide and that fine-tune their activities and/or stability. This review focuses on studies published over the past decade that have provided much insight into the mechanisms of the enzymes that carry out the post-translational modifications.
      >> More from SciFinder ®
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    5. 5
      Kodani, S.; Hudson, M. E.; Durrant, M. C.; Buttner, M. J.; Nodwell, J. R.; Willey, J. M. The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 11448– 11453,  DOI: 10.1073/pnas.0404220101
      5
      The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor
      Kodani, Shinya; Hudson, Michael E.; Durrant, Marcus C.; Buttner, Mark J.; Nodwell, Justin R.; Willey, Joanne M.
      Proceedings of the National Academy of Sciences of the United States of America (2004), 101 (31), 11448-11453CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
      SapB is a morphogenetic peptide that is important for aerial mycelium formation by the filamentous bacterium Streptomyces coelicolor. Prodn. of SapB commences during aerial mycelium formation and depends on most of the genes known to be required for the morphogenesis of aerial hyphae. Furthermore, the application of purified SapB to mutants blocked in morphogenesis restores their capacity to form aerial hyphae. Here, we present evidence that SapB is a lantibiotic-like peptide that is derived by posttranslational modification from the product of a gene (ramS) in the four-gene ram operon, which is under the control of the regulatory gene rams. We show that the product of another gene in the operon (ramC) contains a region that is similar to enzymes involved in the biosynthesis of lantibiotics, suggesting that it might be involved in the posttranslational processing of RamS. We conclude that SapB is derived from RamS through proteolytic cleavage and the introduction of four dehydroalanine residues and two lanthionine bridges. We provide an example of a morphogenetic role for an antibiotic-like mol.
      >> More from SciFinder ®
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    6. 6
      Kodani, S.; Lodato, M. A.; Durrant, M. C.; Picart, F.; Willey, J. M. SapT, a lanthionine-containing peptide involved in aerial hyphae formation in the Streptomycetes. Mol. Microbiol. 2005, 58, 1368– 1380,  DOI: 10.1111/j.1365-2958.2005.04921.x
      6
      SapT, a lanthionine-containing peptide involved in aerial hyphae formation in the streptomycetes
      Kodani, Shinya; Lodato, Michael A.; Durrant, Marcus C.; Picart, Francis; Willey, Joanne M.
      Molecular Microbiology (2005), 58 (5), 1368-1380CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Publishing Ltd.)
      The developmentally complex soil microbe Streptomyces tendae secretes a hydrophobic peptide that restored to developmental mutants of S. coelicolor the ability to raise aerial hyphae. The S. tendae peptide, SapT, has a lantibiotic structure and mol. modeling predicts that it is amphiphilic, making it structurally and functionally similar to the SapB peptide produced by S. coelicolor. However, SapT, which bears three β-Me lanthionine bridges and one lanthionine bridge and demonstrated limited antibiotic activity, is distinct from SapB. The amphiphilic nature of both SapT and SapB is required for their ability to serve as biosurfactants facilitating the emergence of newly formed aerial hyphae. Remarkably, SapB and SapT, and the fungal hydrophobin SC3 were shown to restore to a SapB-deficient S. coelicolor mutant the capacity to undergo complete morphogenesis, such that the extracellular addn. of protein resulted in sporulation. This suggests that the initiation of aerial growth may also indirectly trigger the signal transduction events needed for differentiation. These data imply that the prodn. of morphogenetic peptides may be common among the streptomycetes, but that while their ability to function as biosurfactants is conserved, their specific lantibiotic structure is not. Finally, the identification of a second lanthionine-contg. morphogenetic peptide suggests that lantibiotic structure and function may be more diverse than previously thought.
      >> More from SciFinder ®
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    7. 7
      Willey, J. M.; Willems, A.; Kodani, S.; Nodwell, J. R. Morphogenetic surfactants and their role in the formation of aerial hyphae in Streptomyces coelicolor. Mol. Microbiol. 2006, 59, 731– 742,  DOI: 10.1111/j.1365-2958.2005.05018.x
      7
      Morphogenetic surfactants and their role in the formation of aerial hyphae in Streptomyces coelicolor
      Willey, Joanne M.; Willems, Andrew; Kodani, Shinya; Nodwell, Justin R.
      Molecular Microbiology (2006), 59 (3), 731-742CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Publishing Ltd.)
      A review. Withstanding environmental adversity and seeking optimal conditions for reprodn. are basic requirements for the survival of all organisms. Filamentous bacteria of the genus Streptomyces produce a remarkable cell type called the aerial hyphae that is central to its ability to meet both of these challenges. Recent advances have brought about a major shift in our understanding of the cell surface proteins that play important roles in the generation of these cells. Here we review our current understanding of one of these groups of proteins, the morphogenetic surfactants, with emphasis on the SapB protein of Streptomyces coelicolor.
      >> More from SciFinder ®
      https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28Xit1SgtLg%253D&md5=e362b3d2d36c1ba3d1b3aa0e02e560c9
    8. 8
      Férir, G.; Petrova, M. I.; Andrei, G.; Huskens, D.; Hoorelbeke, B.; Snoeck, R.; Vanderleyden, J.; Balzarini, J.; Bartoschek, S.; Brönstrup, M. The lantibiotic peptide labyrinthopeptin A1 demonstrates broad anti-HIV and anti-HSV activity with potential for microbicidal applications. PLoS One 2013, 8, e64010  DOI: 10.1371/journal.pone.0064010
      8
      The lantibiotic peptide labyrinthopeptin A1 demonstrates broad anti-HIV and anti-HSV activity with potential for microbicidal applications
      Ferir, Geoffrey; Petrova, Mariya I.; Andrei, Graciela; Huskens, Dana; Hoorelbeke, Bart; Snoeck, Robert; Vanderleyden, Jos; Balzarini, Jan; Bartoschek, Stefan; Broenstrup, Mark; Suessmuth, Roderich D.; Schols, Dominique
      PLoS One (2013), 8 (5), e64010CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)
      Lantibiotics are peptides, produced by bacteria, that contain the noncanonical amino acid lanthionine and many of them exhibit antibacterial activities. The labyrinthopeptin A1 (LabyA1) is a prototype peptide of a novel class of carbacyclic lantibiotics. Here, we extensively evaluated its broad-spectrum activity against HIV and HSV in vitro, studied its mechanism of action and evaluated potential microbicidal applications. LabyA1 exhibited a consistent and broad anti-HIV activity (EC50s: 0.70-3.3 μM) and anti-HSV activity (EC50s: 0.29-2.8 μM) in cell cultures. LabyA1 also inhibited viral cell-cell transmission between persistently HIV-infected T cells and uninfected CD4+ T cells (EC50: 2.5 μM) and inhibited the transmission of HIV captured by DC-SIGN+-cells to uninfected CD4+ T cells (EC50: 4.1 μM). Time-of-drug addn. studies revealed that LabyA1 acts as an entry inhibitor against HIV and HSV. Cellular and virus binding studies combined with SPR/FLIPR technol. showed that LabyA1 interacted with the HIV envelope protein gp120, but not with the HIV cellular receptors. LabyA1 also demonstrated additive to synergistic effects in its anti-HIV-1 and anti-HSV-2 activity with anti(retro)viral drugs in dual combinations such as tenofovir, acyclovir, saquinavir, raltegravir and enfuvirtide. LabyA1 can be considered as a novel lead peptide as it had profound antiviral activity against HIV and HSV. Pre-treatment of PBMCs with LabyA1 neither increased the expression of the activation markers CD69 and CD25, nor enhanced HIV replication, nor significantly induced various inflammatory cytokines/chemokines. LabyA1 also did not affect the growth of vaginal Lactobacilli populations. Based on the lack of toxicity on the vaginal Lactobacillus strains and its synergistic/additive profile in combination with clin. approved anti(retro)virals, it deserves further attention as a potential microbicide candidate in the prevention of sexual transmitted diseases.
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    9. 9
      Mohr, K. I.; Volz, C.; Jansen, R.; Wray, V.; Hoffmann, J.; Bernecker, S.; Wink, J.; Gerth, K.; Stadler, M.; Müller, R. Pinensins: the first antifungal lantibiotics. Angew. Chem., Int. Ed. 2015, 54, 11254– 11258,  DOI: 10.1002/anie.201500927
      9
      Pinensins: The first antifungal lantibiotics
      Mohr, Kathrin I.; Volz, Carsten; Jansen, Rolf; Wray, Victor; Hoffmann, Judith; Bernecker, Steffen; Wink, Joachim; Gerth, Klaus; Stadler, Marc; Mueller, Rolf
      Angewandte Chemie, International Edition (2015), 54 (38), 11254-11258CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)
      Lantibiotics (lanthionine-contg. antibiotics) from Gram-pos. bacteria typically exhibit activity against Gram-pos. bacteria. The activity and structure of pinensin A (1) and B (2), lantibiotics isolated from a native Gram-neg. producer Chitinophaga pinensis are described. Surprisingly, the pinensins were found to be highly active against many filamentous fungi and yeasts but show only weak antibacterial activity. To the best of our knowledge, lantibiotic fungicides have not been described before. An in-depth bioinformatic anal. of the biosynthetic gene cluster established the ribosomal origin of these compds. and identified candidate genes encoding all of the enzymes required for post-translational modification. Addnl. encoded functions enabled us to build up a hypothesis for the biosynthesis, export, sensing, and import of this intriguing lantibiotic.
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    10. 10
      Smith, T. E.; Pond, C. D.; Pierce, E.; Harmer, Z. P.; Kwan, J.; Zachariah, M. M.; Harper, M. K.; Wyche, T. P.; Matainaho, T. K.; Bugni, T. S. Accessing chemical diversity from the uncultivated symbionts of small marine animals. Nat. Chem. Biol. 2018, 14, 179– 185,  DOI: 10.1038/nchembio.2537
      10
      Accessing chemical diversity from the uncultivated symbionts of small marine animals
      Smith, Thomas E.; Pond, Christopher D.; Pierce, Elizabeth; Harmer, Zachary P.; Kwan, Jason; Zachariah, Malcolm M.; Harper, Mary Kay; Wyche, Thomas P.; Matainaho, Teatulohi K.; Bugni, Tim S.; Barrows, Louis R.; Ireland, Chris M.; Schmidt, Eric W.
      Nature Chemical Biology (2018), 14 (2), 179-185CODEN: NCBABT; ISSN:1552-4450. (Nature Research)
      Chem. drives many biol. interactions between the microbiota and host animals, yet it is often challenging to identify the chems. involved. This poses a problem, as such small mols. are excellent sources of potential pharmaceuticals, pretested by nature for animal compatibility. We discovered anti-HIV compds. from small, marine tunicates from the Eastern Fields of Papua New Guinea. Tunicates are a reservoir for new bioactive chems., yet their small size often impedes identification or even detection of the chems. within. We solved this problem by combining chem., metagenomics, and synthetic biol. to directly identify and synthesize the natural products. We show that these anti-HIV compds., the divamides, are a novel family of lanthipeptides produced by symbiotic bacteria living in the tunicate. Neighboring animal colonies contain structurally related divamides that differ starkly in their biol. properties, suggesting a role for biosynthetic plasticity in a native context wherein biol. interactions take place.
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    11. 11
      Garg, N.; Salazar-Ocampo, L. M. A.; van der Donk, W. A. In vitro activity of the nisin dehydratase NisB. Proc. Natl. Acad. Sci. U.S.A. 2013, 110, 7258– 7263,  DOI: 10.1073/pnas.1222488110
      11
      In vitro activity of the nisin dehydratase NisB
      Garg, Neha; Salazar-Ocampo, Luis M. A.; van der Donk, Wilfred A.
      Proceedings of the National Academy of Sciences of the United States of America (2013), 110 (18), 7258-7263, S7258/1-S7258/11CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
      The biosynthesis of several classes of ribosomally synthesized and posttranslationally modified peptides involves dehydration of serine and threonine residues. For class I lantibiotics, thiopeptides, and goadsporin, this dehydration is catalyzed by lanthionine biosynthetic enzyme B (LanB) or LanB-like proteins. Although LanB proteins have been studied since 1992, in vitro reconstitution of their dehydration activity has been elusive. We show here the in vitro activity of the dehydratase involved in the biosynthesis of the food preservative nisin (NisB). In vitro, NisB dehydrated its substrate peptide NisA eight times in the presence of glutamate, ATP, Mg2+, and the ribosomal/membrane fraction of bacterial cell ext. Mutation of 23 highly conserved residues of NisB identified a no. of amino acids that are essential for dehydration activity. In addn., these mutagenesis studies identified three mutants, R786A, R826A, and H961A, that result in multiple glutamylations of the NisA substrate. Glutamylation was obsd. during both Escherichia coli coexpression of NisA with these mutants and in vitro assays. Treatment of the glutamylated substrate with WT NisB results in dehydrated NisA, suggesting that the glutamylated peptide is an intermediate in dehydration. Collectively, these studies suggest that dehydration involves glutamylation of the side chains of Ser and Thr followed by elimination. The latter step has precedent in the virginiamycin resistance protein virginiamycin B lyase. These studies will facilitate investigation of other LanB proteins involved in the biosynthesis of lantibiotics, thiopeptides, and goadsporin.
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    12. 12
      Ortega, M. A.; Hao, Y.; Zhang, Q.; Walker, M. C.; van der Donk, W. A.; Nair, S. K. Structure and mechanism of the tRNA-dependent lantibiotic dehydratase NisB. Nature 2015, 517, 509– 512,  DOI: 10.1038/nature13888
      12
      Structure and mechanism of the tRNA-dependent lantibiotic dehydratase NisB
      Ortega, Manuel A.; Hao, Yue; Zhang, Qi; Walker, Mark C.; van der Donk, Wilfred A.; Nair, Satish K.
      Nature (London, United Kingdom) (2015), 517 (7535), 509-512CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)
      Lantibiotics are a class of peptide antibiotics that contain one or more thioether bonds. The lantibiotic, nisin, is an antimicrobial peptide that is widely used as a food preservative to combat food-borne pathogens. Nisin contains dehydroalanine and dehydrobutyrine residues that are formed by the dehydration of Ser/Thr by lantibiotic dehydratase NisB. Recent biochem. studies revealed that NisB glutamylates Ser/Thr side-chains as part of the dehydration process. However, the mol. mechanism by which NisB uses glutamate to catalyze dehydration remains unresolved. Here, the authors show that this process involves glutamyl-tRNAGlu to activate the Ser/Thr residues. In addn., the 2.9-Å crystal structure of Escherichia coli NisB in complex with its substrate peptide, nisin A, reveals the presence of 2 sep. domains that catalyze the Ser/Thr glutamylation and glutamate elimination steps. The co-crystal structure also provided insights into substrate recognition by lantibiotic dehydratases. These findings demonstrated an unexpected role for aminoacyl-tRNA in the formation of dehydroamino acids in lantibiotics, and served as a basis for the functional characterization of the many lantibiotic-like dehydratases involved in the biosynthesis of other classes of natural products.
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    13. 13
      Ortega, M. A.; Hao, Y.; Walker, M. C.; Donadio, S.; Sosio, M.; Nair, S. K.; van der Donk, W. A. Structure and tRNA specificity of MibB, a lantibiotic dehydratase from Actinobacteria involved in NAI-107 biosynthesis. Cell Chem. Biol. 2016, 23, 370– 380,  DOI: 10.1016/j.chembiol.2015.11.017
      13
      Structure and tRNA Specificity of MibB, a Lantibiotic Dehydratase from Actinobacteria Involved in NAI-107 Biosynthesis
      Ortega Manuel A; Hao Yue; Nair Satish K; van der Donk Wilfred A; Walker Mark C; van der Donk Wilfred A; Donadio Stefano; Sosio Margherita; Nair Satish K
      Cell chemical biology (2016), 23 (3), 370-380 ISSN:.
      Class I lantibiotic dehydratases dehydrate selected Ser/Thr residues of a precursor peptide. Recent studies demonstrated the requirement of glutamyl-tRNA(Glu) for Ser/Thr activation by one of these enzymes (NisB) from the Firmicute Lactococcus lactis. However, the generality of glutamyl-tRNA(Glu) usage and the tRNA specificity of lantibiotic dehydratases have not been established. Here we report the 2.7-ÅA resolution crystal structure, along with the glutamyl-tRNA(Glu) utilization of MibB, a lantibiotic dehydratase from the Actinobacterium Microbispora sp. 107891 involved in the biosynthesis of the clinical candidate NAI-107. Biochemical assays revealed nucleotides A73 and U72 within the tRNA(Glu) acceptor stem to be important for MibB glutamyl-tRNA(Glu) usage. Using this knowledge, an expression system for the production of NAI-107 analogs in Escherichia coli was developed, overcoming the inability of MibB to utilize E. coli tRNA(Glu). Our work provides evidence for a common tRNA(Glu)-dependent dehydration mechanism, paving the way for the characterization of lantibiotics from various phyla.
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    14. 14
      Bothwell, I. R.; Cogan, D. P.; Kim, T.; Reinhardt, C. J.; van der Donk, W. A.; Nair, S. K. Characterization of glutamyl-tRNA-dependent dehydratases using nonreactive substrate mimics. Proc. Natl. Acad. Sci. U.S.A. 2019, 116, 17245– 17250,  DOI: 10.1073/pnas.1905240116
      14
      Characterization of glutamyl-tRNA-dependent dehydratases using nonreactive substrate mimics
      Bothwell, Ian R.; Cogan, Dillon P.; Kim, Terry; Reinhardt, Christopher J.; van der Donk, Wilfred A.; Nair, Satish K.
      Proceedings of the National Academy of Sciences of the United States of America (2019), 116 (35), 17245-17250CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
      The peptide natural product nisin has been used as a food preservative for 6 decades with minimal development of resistance. Nisin contains the unusual amino acids dehydroalanine and dehydrobutyrine, which are posttranslationally installed by class I lanthipeptide dehydratases (LanBs) on a linear peptide substrate through an unusual glutamyl-tRNA-dependent dehydration of Ser and Thr. To date, little is known about how LanBs catalyze the transfer of glutamate from charged tRNAGlu to the peptide substrate, or how they carry out the subsequent elimination of the peptide-glutamyl adducts to afford dehydro amino acids. Here, we describe the synthesis of inert analogs that mimic substrate glutamyl-tRNAGlu and the glutamylated peptide intermediate, and det. the crystal structures of 2 LanBs in complex with each of these compds. Mutational studies were used to characterize the function of the glutamylation and glutamate elimination active-site residues identified through the structural anal. These combined studies provide insights into the mechanisms of substrate recognition, glutamylation, and glutamate elimination by LanBs to effect a net dehydration reaction of Ser and Thr.
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    15. 15
      Li, B.; Yu, J. P. J.; Brunzelle, J. S.; Moll, G. N.; van der Donk, W. A.; Nair, S. K. Structure and mechanism of the lantibiotic cyclase involved in nisin biosynthesis. Science 2006, 311, 1464– 1467,  DOI: 10.1126/science.1121422
      15
      Structure and Mechanism of the Lantibiotic Cyclase Involved in Nisin Biosynthesis
      Li, Bo; Yu, John Paul J.; Brunzelle, Joseph S.; Moll, Gert N.; van der Donk, Wilfred A.; Nair, Satish K.
      Science (Washington, DC, United States) (2006), 311 (5766), 1464-1467CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)
      Nisin is a posttranslationally modified antimicrobial peptide that is widely used as a food preservative. It contains five cyclic thioethers of varying sizes that are installed by a single enzyme, NisC. Reported here are the in vitro reconstitution of the cyclization process and the x-ray crystal structure of the NisC enzyme. The structure reveals similarities in fold and substrate activation with mammalian farnesyl transferases, suggesting that human homologs of NisC posttranslationally modify a cysteine of a protein substrate.
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    16. 16
      Li, B.; van der Donk, W. A. Identification of essential catalytic residues of the cyclase NisC involved in the biosynthesis of nisin. J. Biol. Chem. 2007, 282, 21169– 21175,  DOI: 10.1074/jbc.M701802200
      16
      Identification of Essential Catalytic Residues of the Cyclase NisC Involved in the Biosynthesis of Nisin
      Li, Bo; van der Donk, Wilfred A.
      Journal of Biological Chemistry (2007), 282 (29), 21169-21175CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)
      Nisin is a post-translationally modified antimicrobial peptide that has been widely used in the food industry for several decades. It contains five cyclic thioether cross-links of varying sizes that are installed by a single enzyme, NisC, that catalyzes the addn. of cysteines to dehydroamino acids. The recent X-ray crystal structure of NisC has provided the first insights into the catalytic residues responsible for the cyclization step during nisin biosynthesis. In this study, the conserved residues His212, Arg280, Asp141, and Tyr285 as well as the ligands to the zinc in the active site (Cys284, Cys330, and His331) were substituted by site-directed mutagenesis. Binding studies showed that all mutants had similar affinities for NisA. Activity assays showed that whereas His212 and Asp141 were essential for correct cyclization as judged by the antimicrobial activity of the final product, Arg280 and Tyr285 were not. Mutation of zinc ligands to alanine also abolished the enzymic activity, and these mutant proteins were shown to contain decreased levels of zinc. These results show that the zinc is essential for activity and support a model in which the zinc is used to activate the cysteines in the substrate for nucleophilic attack. These findings also argue against an essential role of Arg280 and Tyr285 as an active site general acid/base in the mechanism of cyclization.
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    17. 17
      Mukherjee, S.; Huo, L.; Thibodeaux, G. N.; van der Donk, W. A. Synthesis and bioactivity of diastereomers of the virulence lanthipeptide cytolysin. Org. Lett. 2016, 18, 6188– 6191,  DOI: 10.1021/acs.orglett.6b03246
      17
      Synthesis and bioactivity of diastereomers of the virulence lanthipeptide cytolysin
      Mukherjee, Subha; Huo, Liujie; Thibodeaux, Gabrielle N.; van der Donk, Wilfred A.
      Organic Letters (2016), 18 (23), 6188-6191CODEN: ORLEF7; ISSN:1523-7052. (American Chemical Society)
      Cytolysin, a two-component lanthipeptide comprising cytolysin S (CylLS'') and cytolysin L (CylLL''), is the only family member to exhibit lytic activity against mammalian cells in addn. to synergistic antimicrobial activity. A subset of the thioether cross-links of CylLS'' and CylLL'' have LL stereochem. instead of the canonical DL stereochem. in all previously characterized lanthipeptides. The synthesis of a CylLS'' variant with DL stereochem. is reported. Its antimicrobial activity was found to be decreased, but not its lytic activity against red blood cells. Hence, the unusual LL stereochem. is not responsible for the lytic activity.
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    18. 18
      Knerr, P. J.; van der Donk, W. A. Chemical synthesis of the lantibiotic lacticin 481 reveals the importance of lanthionine stereochemistry. J. Am. Chem. Soc. 2013, 135, 7094– 7097,  DOI: 10.1021/ja4014024
      18
      Chemical synthesis of the lantibiotic Lacticin 481 reveals the importance of lanthionine stereochemistry
      Knerr, Patrick J.; van der Donk, Wilfred A.
      Journal of the American Chemical Society (2013), 135 (19), 7094-7097CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lantibiotics are a family of antibacterial peptide natural products characterized by the post-translational installation of the thioether-contg. amino acids lanthionine and methyllanthionine. Until recently, only a single naturally occurring stereochem. configuration for each of these cross-links was known. The discovery of lantibiotics with alternative lanthionine and methyllanthionine stereochem. has prompted an investigation of its importance to biol. activity. Here, solid-supported chem. synthesis enabled the total synthesis of the lantibiotic lacticin 481 and analogs contg. cross-links with non-native stereochem. configurations. Biol. evaluation revealed that these alterations abolished the antibacterial activity in all of the analogs, revealing the crit. importance of the enzymically installed stereochem. for the biol. activity of lacticin 481.
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    19. 19
      Tang, W.; van der Donk, W. A. The sequence of the enterococcal cytolysin imparts unusual lanthionine stereochemistry. Nat. Chem. Biol. 2013, 9, 157– 159,  DOI: 10.1038/nchembio.1162
      19
      The sequence of the enterococcal cytolysin imparts unusual lanthionine stereochemistry
      Tang, Weixin; van der Donk, Wilfred A.
      Nature Chemical Biology (2013), 9 (3), 157-159CODEN: NCBABT; ISSN:1552-4450. (Nature Publishing Group)
      The enterococcal cytolysin is a two-component lantibiotic of unknown structure with hemolytic activity that is important for virulence. We prepd. cytolysin by coexpression of each precursor peptide with the synthetase CylM in Escherichia coli and characterized its structure. Unexpectedly, cytolysin is to our knowledge the first example of a lantibiotic contg. lanthionine and methyllanthionine structures with different stereochemistries in the same peptide. The stereochem. is detd. by the sequence of the substrate peptide.
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    20. 20
      Tang, W.; Jiménez-Osés, G.; Houk, K. N.; van der Donk, W. A. Substrate control in stereoselective lanthionine biosynthesis. Nat. Chem. 2015, 7, 57– 64,  DOI: 10.1038/nchem.2113
      20
      Substrate control in stereoselective lanthionine biosynthesis
      Tang, Weixin; Jimenez-Oses, Gonzalo; Houk, K. N.; van der Donk, Wilfred A.
      Nature Chemistry (2015), 7 (1), 57-64CODEN: NCAHBB; ISSN:1755-4330. (Nature Publishing Group)
      Enzymes are typically highly stereoselective catalysts that enforce a reactive conformation on their native substrates. We report here a rare example in which the substrate controls the stereoselectivity of an enzyme-catalyzed Michael-type addn. during the biosynthesis of lanthipeptides. These natural products contain thioether crosslinks formed by a cysteine attack on dehydrated Ser and Thr residues. We demonstrate that several lanthionine synthetases catalyze highly selective anti-addns. in which the substrate (and not the enzyme) dets. whether the addn. occurs from the re or si face. A single point mutation in the peptide substrate completely inverted the stereochem. outcome of the enzymic modification. Quantum mech. calcns. reproduced the exptl. obsd. selectivity and suggest that conformational restraints imposed by the amino-acid sequence on the transition states det. the face selectivity of the Michael-type cyclization.
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    21. 21
      Tang, W.; Thibodeaux, G. N.; van der Donk, W. A. The enterococcal cytolysin synthetase coevolves with substrate for stereoselective lanthionine synthesis. ACS Chem. Biol. 2016, 11, 2438– 2446,  DOI: 10.1021/acschembio.6b00397
      21
      The enterococcal cytolysin synthetase coevolves with substrate for stereoselective lanthionine synthesis
      Tang, Weixin; Thibodeaux, Gabrielle N.; van der Donk, Wilfred A.
      ACS Chemical Biology (2016), 11 (9), 2438-2446CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)
      Stereochem. control is crit. in natural product biosynthesis. For ribosomally synthesized and post-translationally modified peptides (RiPPs), the mechanism(s) by which stereoselectivity is achieved is still poorly understood. In this work, we focused on the stereoselective lanthionine synthesis in lanthipeptides, a major class of RiPPs formed by the addn. of Cys residues to dehydroalanine (Dha) or dehydrobutyrine (Dhb). Nonenzymic cyclization of the small subunit of a virulence lanthipeptide, the enterococcal cytolysin, resulted in the native modified peptide as the major product, suggesting that both regioselectivity and stereoselectivity are inherent to the dehydrated peptide sequence. These results support previous computational studies that a Dhx-Dhx-Xxx-Xxx-Cys motif (Dhx = Dha or Dhb; Xxx = any amino acid except Dha, Dhb, and Cys) preferentially cyclizes by attack on the Re face of Dha or Dhb. Characterization of the stereochem. of the products formed enzymically with substrate mutants revealed that the lanthionine synthetase actively reinforces Re face attack. These findings support the hypothesis of substrate-controlled selectivity in lanthionine synthesis but also reveal likely coevolution of substrates and lanthionine synthetases to ensure the stereoselective synthesis of lanthipeptides with defined biol. activities.
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    22. 22
      Garg, N.; Goto, Y.; Chen, T.; van der Donk, W. A. Characterization of the stereochemical configuration of lanthionines formed by the lanthipeptide synthetase GeoM. Biopolymers 2016, 106, 834– 842,  DOI: 10.1002/bip.22876
      22
      Characterization of the stereochemical configuration of lanthionines formed by the lanthipeptide synthetase GeoM
      Garg, Neha; Goto, Yuki; Chen, Ting; van der Donk, Wilfred A.
      Biopolymers (2016), 106 (6), 834-842CODEN: BIPMAA; ISSN:0006-3525. (John Wiley & Sons, Inc.)
      The biosynthesis of the class II lanthipeptide geobacillin II was reconstituted in vitro. The purified precursor peptide was modified by the lanthipeptide synthetase GeoM at temps. ranging between 37 and 80°C demonstrating the thermostability of the enzyme. Geobacillin II shares with cytolysin, haloduracin, and carnolysin a DhxDhxXxxXxxCys motif (Dhx = dehydroalanine or dehydrobutyrine) as precursor to its N-terminal A-ring. Like in these other three lantibiotics, the lanthionine in the A-ring of geobacillin II had the LL stereochem. configuration as shown by chiral gas chromatog./mass spectrometry. Various analogs of geobacillin II were produced using co-expression of mutants of the precursor peptide GeoAII and the synthetase GeoM in Escherichia coli. The findings in this study suggest that the stereochem. outcome of the A-ring in geobacillin II is not solely dependent on the peptide sequence as previously suggested for haloduracin © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 834-842, 2016.
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    23. 23
      Lohans, C. T.; Li, J. L.; Vederas, J. C. Structure and biosynthesis of carnolysin, a homologue of enterococcal cytolysin with D-amino acids. J. Am. Chem. Soc. 2014, 136, 13150– 13153,  DOI: 10.1021/ja5070813
      23
      Structure and biosynthesis of carnolysin, a homolog of enterococcal cytolysin with D-amino acids
      Lohans, Christopher T.; Li, Jessica L.; Vederas, John C.
      Journal of the American Chemical Society (2014), 136 (38), 13150-13153CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lantibiotics are a group of highly post-translationally modified bacterial antimicrobial peptides characterized by the presence of the thioether-contg. amino acids lanthionine and methyllanthionine. Carnobacterium maltaromaticum C2 was found to produce a two-component lantibiotic homologous to enterococcal cytolysin. Through tandem mass spectrometry and NMR spectroscopy, the post-translational modifications of carnolysin were established, and the topologies of the lanthionine and methyllanthionine rings were detd. Chiral GC-MS anal. revealed that, like cytolysin, carnolysin contained lanthionine and methyllanthionine residues of unusual stereochem. Carnolysin, unlike cytolysin, was shown to contain D-alanine and unprecedented D-aminobutyrate derived from serine and threonine, resp. Carnolysin was heterologously expressed in Escherichia coli, demonstrating that reductase CrnJ is involved in the formation of the D-amino acids.
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    24. 24
      Sarksian, R.; Hegemann, J. D.; Simon, M. A.; Acedo, J. Z.; van der Donk, W. A. Unexpected methyllanthionine stereochemistry in the morphogenetic lanthipeptide SapT. J. Am. Chem. Soc. 2022, 144, 6373– 6382,  DOI: 10.1021/jacs.2c00517
      24
      Unexpected methyllanthionine stereochemistry in the morphogenetic lanthipeptide SapT
      Sarksian, Raymond; Hegemann, Julian D.; Simon, Max A.; Acedo, Jeella Z.; van der Donk, Wilfred A.
      Journal of the American Chemical Society (2022), 144 (14), 6373-6382CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lanthipeptides are polycyclic peptides characterized by the presence of lanthionine (Lan) and/or methyllanthionine (MeLan). They are members of the ribosomally synthesized and post-translationally modified peptides (RiPPs). The stereochem. configuration of (Me)Lan crosslinks is important for the bioactivity of lanthipeptides. To date, MeLan residues in characterized lanthipeptides have either the 2S,3S or 2R,3R stereochem. Herein, we reconstituted in Escherichia coli the biosynthetic pathway toward SapT, a class I lanthipeptide that exhibits morphogenetic activity. Through the synthesis of stds., the heterologously produced peptide was shown to possess three MeLan residues with the 2S,3R stereochem. (D-allo-L-MeLan), the first time such stereochem. has been obsd. in a lanthipeptide. Bioinformatic anal. of the biosynthetic enzymes suggests this stereochem. may also be present in other lanthipeptides. Anal. of another gene cluster in Streptomyces coelicolor that is widespread in actinobacteria confirmed another example of D-allo-L-MeLan and verified the bioinformatic prediction. We propose a mechanism for the origin of the unexpected stereochem. and provide support using site-directed mutagenesis.
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    25. 25
      Bothwell, I. R.; Caetano, T.; Sarksian, R.; Mendo, S.; van der Donk, W. A. Structural analysis of class I lanthipeptides from Pedobacter lusitanus NL19 reveals an unusual ring pattern. ACS Chem. Biol. 2021, 16, 1019– 1029,  DOI: 10.1021/acschembio.1c00106
      25
      Structural Analysis of Class I Lanthipeptides from Pedobacter lusitanus NL19 Reveals an Unusual Ring Pattern
      Bothwell, Ian R.; Caetano, Tania; Sarksian, Raymond; Mendo, Sonia; van der Donk, Wilfred A.
      ACS Chemical Biology (2021), 16 (6), 1019-1029CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)
      Lanthipeptides are ribosomally synthesized and post-translationally modified peptide natural products characterized by the presence of lanthionine and methyllanthionine crosslinked amino acids formed by dehydration of Ser/Thr residues followed by conjugate addn. of Cys to the resulting dehydroamino acids. Class I lanthipeptide dehydratases use glutamyl-tRNAGlu as a cosubstrate to glutamylate Ser/Thr followed by glutamate elimination. A vast majority of lanthipeptides identified from class I synthase systems were from Gram-pos. bacteria. Herein, the authors report the heterologous expression and modification in Escherichia coli of two lanthipeptides from the Gram-neg. Bacteroidetes Pedobacter lusitanus NL19. These peptides are representative of a group of compds. frequently encoded in Pedobacter genomes. Structural characterization of the lanthipeptides revealed a novel ring pattern as well as an unusual LL-lanthionine stereochem. configuration and a cyclase that lacks the canonical zinc ligands found in most LanC enzymes.
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    26. 26
      Acedo, J. Z.; Bothwell, I. R.; An, L.; Trouth, A.; Frazier, C.; van der Donk, W. A. O-methyltransferase-mediated incorporation of a β-amino acid in lanthipeptides. J. Am. Chem. Soc. 2019, 141, 16790– 16801,  DOI: 10.1021/jacs.9b07396
      26
      O-Methyltransferase-Mediated Incorporation of a β-Amino Acid in Lanthipeptides
      Acedo, Jeella Z.; Bothwell, Ian R.; An, Linna; Trouth, Abby; Frazier, Clara; van der Donk, Wilfred A.
      Journal of the American Chemical Society (2019), 141 (42), 16790-16801CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lanthipeptides represent a large class of cyclic natural products defined by the presence of lanthionine (Lan) and methyllanthionine (MeLan) cross-links. With the advances in DNA sequencing technologies and genome mining tools, new biosynthetic enzymes capable of installing unusual structural features are continuously being discovered. In this study, we investigated an O-methyltransferase that is a member of the most prominent auxiliary enzyme family assocd. with class I lanthipeptide biosynthetic gene clusters. Despite the prevalence of these enzymes, their function has not been established. Herein, we demonstrate that the O-methyltransferase OlvSA encoded in the olv gene cluster from Streptomyces olivaceus NRRL B-3009 catalyzes the rearrangement of a highly conserved aspartate residue to a β-amino acid, isoaspartate, in the lanthipeptide OlvA(BCSA). We elucidated the NMR soln. structure of the GluC-digested peptide, OlvA(BCSA)GluC, which revealed a unique ring topol. comprising four interlocking rings and positions the isoaspartate residue in a solvent exposed loop that is stabilized by a MeLan ring. Gas chromatog.-mass spectrometry anal. further indicated that OlvA(BCSA) contains two DL-MeLan rings and two Lan rings with an unusual LL-stereochem. Lastly, in vitro reconstitution of OlvSA activity showed that it is a leader peptide-independent and S-adenosyl methionine-dependent O-methyltransferase that mediates the conversion of a highly conserved aspartate residue in a cyclic substrate into a succinimide, which is hydrolyzed to generate an Asp or isoAsp contg. peptide. This overall transformation converts an α-amino acid into a β-amino acid in a ribosomally synthesized peptide, via an electrophilic intermediate that may be the intended product.
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    27. 27
      Marsh, A. J.; O’Sullivan, O.; Ross, R. P.; Cotter, P. D.; Hill, C. In silico analysis highlights the frequency and diversity of type 1 lantibiotic gene clusters in genome sequenced bacteria. BMC Genomics 2010, 11, 679  DOI: 10.1186/1471-2164-11-679
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      In silico analysis highlights the frequency and diversity of type 1 lantibiotic gene clusters in genome sequenced bacteria
      Marsh, Alan J.; O'Sullivan, Orla; Ross, R. Paul; Cotter, Paul D.; Hill, Colin
      BMC Genomics (2010), 11 (), 679CODEN: BGMEET; ISSN:1471-2164. (BioMed Central Ltd.)
      Background: Lantibiotics are lanthionine-contg., post-translationally modified antimicrobial peptides. These peptides have significant, but largely untapped, potential as preservatives and chemotherapeutic agents. Type 1 lantibiotics are those in which lanthionine residues are introduced into the structural peptide (LanA) through the activity of sep. lanthionine dehydratase (LanB) and lanthionine synthetase (LanC) enzymes. Here we take advantage of the conserved nature of LanC enzymes to devise an in silico approach to identify potential lantibiotic-encoding gene clusters in genome sequenced bacteria. Results: In total 49 novel type 1 lantibiotic clusters were identified which unexpectedly were assocd. with species, genera and even phyla of bacteria which have not previously been assocd. with lantibiotic prodn. Conclusions: Multiple type 1 lantibiotic gene clusters were identified at a frequency that suggests that these antimicrobials are much more widespread than previously thought. These clusters represent a rich repository which can yield a large no. of valuable novel antimicrobials and biosynthetic enzymes.
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    28. 28
      Walker, M. C.; Eslami, S. M.; Hetrick, K. J.; Ackenhusen, S. E.; Mitchell, D. A.; van der Donk, W. A. Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family. BMC Genomics 2020, 21, 387  DOI: 10.1186/s12864-020-06785-7
      28
      Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family
      Walker, Mark C.; Eslami, Sara M.; Hetrick, Kenton J.; Ackenhusen, Sarah E.; Mitchell, Douglas A.; van der Donk, Wilfred A.
      BMC Genomics (2020), 21 (1), 387CODEN: BGMEET; ISSN:1471-2164. (BioMed Central Ltd.)
      Lanthipeptides belong to the ribosomally synthesized and post-translationally modified peptide group of natural products and have a variety of biol. activities ranging from antibiotics to antinociceptives. These peptides are cyclized through thioether crosslinks and can bear other secondary post-translational modifications. While lanthipeptide biosynthetic gene clusters can be identified by the presence of genes encoding characteristic enzymes involved in the post-translational modification process, locating the precursor peptides encoded within these clusters is challenging due to their short length and high sequence variability, which limits the high-throughput exploration of lanthipeptide biosynthesis. To address this challenge, we enhanced the predictive capabilities of Rapid ORF Description & Evaluation Online (RODEO) to identify members of all four known classes of lanthipeptides. Using RODEO, we mined over 100,000 bacterial and archaeal genomes in the RefSeq database. We identified nearly 8500 lanthipeptide precursor peptides. These precursor peptides were identified in a broad range of bacterial phyla as well as the Euryarchaeota phylum of archaea. Bacteroidetes were found to encode a large no. of these biosynthetic gene clusters, despite making up a relatively small portion of the genomes in this dataset. A no. of these precursor peptides are similar to those of previously characterized lanthipeptides, but even more were not, including potential antibiotics. One such new antimicrobial lanthipeptide was purified and characterized. Addnl., examn. of the biosynthetic gene clusters revealed that enzymes installing secondary post-translational modifications are more widespread than initially thought. Lanthipeptide biosynthetic gene clusters are more widely distributed and the precursor peptides encoded within these clusters are more diverse than previously appreciated, demonstrating that the lanthipeptide sequence-function space remains largely underexplored.
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    29. 29
      Bentley, S. D.; Chater, K. F.; Cerdeno-Tarraga, A. M.; Challis, G. L.; Thomson, N. R.; James, K. D.; Harris, D. E.; Quail, M. A.; Kieser, H.; Harper, D. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature 2002, 417, 141– 147,  DOI: 10.1038/417141a
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      Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2)
      Bentley, S. D.; Chater, K. F.; Cerdeno-Tarraga, A.-M.; Challis, G. L.; Thomson, N. R.; James, K. D.; Harris, D. E.; Quail, M. A.; Kieser, H.; Harper, D.; Bateman, A.; Brown, S.; Chandra, G.; Chen, C. W.; Collins, M.; Cronin, A.; Fraser, A.; Goble, A.; Hidalgo, J.; Hornsby, T.; Howarth, S.; Huang, C.-H.; Kieser, T.; Larke, L.; Murphy, L.; Oliver, K.; O'Neil, S.; Rabbinowitsch, E.; Rajandream, M.-A.; Rutherford, K.; Rutter, S.; Seeger, K.; Saunders, D.; Sharp, S.; Squares, R.; Squares, S.; Taylor, K.; Warren, T.; Wietzorrek, A.; Woodward, J.; Barrell, B. G.; Parkhill, J.; Hopwood, D. A.
      Nature (London, United Kingdom) (2002), 417 (6885), 141-147CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)
      Streptomyces coelicolor is a representative of the group of soil-dwelling, filamentous bacteria responsible for producing most natural antibiotics used in human and veterinary medicine. The 8,667,507 base pair linear chromosome of this organism, contg. the largest no. of genes so far discovered in a bacterium, is reported. The 7825 predicted genes include more than 20 clusters coding for known or predicted secondary metabolites. The genome contains an unprecedented proportion of regulatory genes, predominantly those likely to be involved in responses to external stimuli and stresses, and many duplicated gene sets that may represent 'tissue-specific' isoforms operating in different phases of colonial development, a unique situation for a bacterium. An ancient synteny was revealed between the central 'core' of the chromosome and the whole chromosome of pathogens Mycobacterium tuberculosis and Corynebacterium diphtheriae. The genome sequence will greatly increase our understanding of microbial life in the soil as well as aiding the generation of new drug candidates by genetic engineering. The complete sequence is deposited in GenBank/EMBL under accession no. AL645882. [This abstr. record is one of two records for this document necessitated by the large no. of index entries required to fully index the document and publication system constraints.].
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    30. 30
      Challis, G. L. Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways. J. Ind. Microbiol. Biotechnol. 2014, 41, 219– 232,  DOI: 10.1007/s10295-013-1383-2
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      Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways
      Challis, Gregory L.
      Journal of Industrial Microbiology & Biotechnology (2014), 41 (2), 219-232CODEN: JIMBFL; ISSN:1367-5435. (Springer)
      A review. Streptomyces, and related genera of Actinobacteria, are renowned for their ability to produce antibiotics and other bioactive natural products with a wide range of applications in medicine and agriculture. Streptomyces coelicolor A3(2) is a model organism that has been used for more than five decades to study the genetic and biochem. basis for the prodn. of bioactive metabolites. In 2002, the complete genome sequence of S. coelicolor was published. This greatly accelerated progress in understanding the biosynthesis of metabolites known or suspected to be produced by S. coelicolor and revealed that streptomycetes have far greater potential to produce bioactive natural products than suggested by classical bioassay-guided isolation studies. In this article, efforts to exploit the S. coelicolor genome sequence for the discovery of novel natural products and biosynthetic pathways are summarized.
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    31. 31
      Ozaki, T.; Nishiyama, M.; Kuzuyama, T. Novel tryptophan metabolism by a potential gene cluster that is widely distributed among actinomycetes. J. Biol. Chem. 2013, 288, 9946– 9956,  DOI: 10.1074/jbc.M112.436451
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      Novel tryptophan metabolism by a potential gene cluster that is widely distributed among Actinomycetes
      Ozaki, Taro; Nishiyama, Makoto; Kuzuyama, Tomohisa
      Journal of Biological Chemistry (2013), 288 (14), 9946-9956CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)
      The characterization of potential gene clusters is a promising strategy for the identification of novel natural products and the expansion of structural diversity. However, there are often difficulties in identifying potential metabolites because their biosynthetic genes are either silenced or expressed only at a low level. Here, we report the identification of a novel metabolite that is synthesized by a potential gene cluster contg. an indole prenyltransferase gene (SCO7467) and a flavin-dependent monooxygenase (FMO) gene (SCO7468), which were mined from the genome of Streptomyces coelicolor A3(2). We introduced these two genes into the closely related Streptomyces lividans TK23 and analyzed the culture broths of the transformants. This process allowed us to identify a novel metabolite, 5-dimethylallylindole-3-acetonitrile (5-DMAIAN) that was overproduced in the transformant. Biochem. characterization of the recombinant SCO7467 and SCO7468 demonstrated the novel l-tryptophan metab. leading to 5-DMAIAN. SCO7467 catalyzes the prenylation of l-tryptophan to form 5-dimethylallyl-l-tryptophan (5-DMAT). This enzyme is the first actinomycetes prenyltransferase known to catalyze the addn. of a dimethylallyl group to the C-5 of tryptophan. SCO7468 then catalyzes the conversion of 5-DMAT into 5-dimethylallylindole-3-acetaldoxime (5-DMAIAOx). An aldoxime-forming reaction catalyzed by the FMO enzyme was also identified for the first time in this study. Finally, dehydration of 5-DMAIAOx presumably occurs to yield 5-DMAIAN. This study provides insight into the biosynthesis of prenylated indoles that have been purified from actinomycetes.
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    32. 32
      Nodwell, J. R. Microbe profile: Streptomyces coelicolor: a burlesque of pigments and phenotypes. Microbiology 2019, 165, 953– 955,  DOI: 10.1099/mic.0.000821
      32
      Microbe profile: Streptomyces coelicolor: a burlesque of pigments and phenotypes
      Nodwell, Justin R.
      Microbiology (London, United Kingdom) (2019), 165 (9), 953-955CODEN: MROBEO; ISSN:1465-2080. (Microbiology Society)
      The streptomycetes are soil-dwelling bacteria that are found in soil everywhere on Earth: the mol. geosmin, which they produce as part of their life cycle, is what gives soil its familiar 'earthy' smell. The species is best known for the prodn. of biol. active small mols. called 'natural products'. These mols. are the source of most of our antibiotics and anti-fungals, as well as many other drugs. The streptomycetes have a filamentous form rather than the more familiar rod-shaped spirochete and coccoid forms. They exhibit a complex life cycle and sporulation mechanism involving several differentiated cell types, each having specific roles in the colony life history. Streptomyces coelicolor is an important model system for this genus - research on this bacterium has provided foundational information for all of these fascinating processes.
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    33. 33
      Lautru, S.; Deeth, R. J.; Bailey, L. M.; Challis, G. L. Discovery of a new peptide natural product by Streptomyces coelicolor genome mining. Nat. Chem. Biol. 2005, 1, 265– 269,  DOI: 10.1038/nchembio731
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      Discovery of a new peptide natural product by Streptomyces coelicolor genome mining
      Lautru, Sylvie; Deeth, Robert J.; Bailey, Lianne M.; Challis, Gregory L.
      Nature Chemical Biology (2005), 1 (5), 265-269CODEN: NCBABT; ISSN:1552-4450. (Nature Publishing Group)
      Analyses of microbial genome sequences reveal numerous examples of gene clusters encoding proteins typically involved in complex natural product biosynthesis but not assocd. with the prodn. of known natural products. In Streptomyces coelicolor M145 there are several gene clusters encoding new nonribosomal peptide synthetase (NRPS) systems not assocd. with known metabolites. Application of structure-based models for substrate recognition by NRPS adenylation domains predicts the amino acids incorporated into the putative peptide products of these systems, but the accuracy of these predictions is untested. Here we report the isolation and structure detn. of the new tris-hydroxamate tetrapeptide iron chelator coelichelin from S. coelicolor using a genome mining approach guided by substrate predictions for the trimodular NRPS CchH, and we show that this enzyme, which lacks a C-terminal thioesterase domain, together with a homolog of enterobactin esterase (CchJ), are required for coelichelin biosynthesis. These results demonstrate that accurate prediction of adenylation domain substrate selectivity is possible and raise intriguing mechanistic questions regarding the assembly of a tetrapeptide by a trimodular NRPS.
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    34. 34
      Song, L.; Barona-Gomez, F.; Corre, C.; Xiang, L.; Udwary, D. W.; Austin, M. B.; Noel, J. P.; Moore, B. S.; Challis, G. L. Type III polyketide synthase beta-ketoacyl-ACP starter unit and ethylmalonyl-CoA extender unit selectivity discovered by Streptomyces coelicolor genome mining. J. Am. Chem. Soc. 2006, 128, 14754– 14755,  DOI: 10.1021/ja065247w
      34
      Type III Polyketide Synthase β-Ketoacyl-ACP Starter Unit and Ethylmalonyl-CoA Extender Unit Selectivity Discovered by Streptomyces coelicolor Genome Mining
      Song, Lijiang; Barona-Gomez, Francisco; Corre, Christophe; Xiang, Longkuan; Udwary, Daniel W.; Austin, Michael B.; Noel, Joseph P.; Moore, Bradley S.; Challis, Gregory L.
      Journal of the American Chemical Society (2006), 128 (46), 14754-14755CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Polyketide synthases (PKSs) are involved in the biosynthesis of many important natural products. In bacteria, type III PKSs typically catalyze iterative decarboxylation and condensation reactions of malonyl-CoA building blocks in the biosynthesis of polyhydroxyarom. products. Here it is shown that Gcs, a type III PKS encoded by the sco7221 ORF of the bacterium Streptomyces coelicolor, is required for biosynthesis of the germicidin family of 3,6-dialkyl-4-hydroxypyran-2-one natural products. Evidence consistent with Gcs-catalyzed elongation of specific β-ketoacyl-ACP products of the fatty acid synthase FabH with ethyl- or methylmalonyl-CoA in the biosynthesis of germicidins is presented. Selectivity for β-ketoacyl-ACP starter units and ethylmalonyl-CoA as an extender unit is unprecedented for type III PKSs, suggesting these enzymes may be capable of utilizing a far wider range of starter and extender units for natural product assembly than believed until now.
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    35. 35
      Lin, X.; Hopson, R.; Cane, D. E. Genome mining in Streptomyces coelicolor: molecular cloning and characterization of a new sesquiterpene synthase. J. Am. Chem. Soc. 2006, 128, 6022– 6023,  DOI: 10.1021/ja061292s
      35
      Genome Mining in Streptomyces coelicolor: Molecular Cloning and Characterization of a New Sesquiterpene Synthase
      Lin, Xin; Hopson, Russell; Cane, David E.
      Journal of the American Chemical Society (2006), 128 (18), 6022-6023CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      The terpene synthase encoded by the SCO5222 (SC7E4.19) gene of Streptomyces coelicolor was cloned by PCR and expressed in Escherichia coli as an N-terminal-His6-tag protein. Incubation of the recombinant protein, SCO5222p, with farnesyl diphosphate (1, FPP) in the presence of Mg(II) gave a new sesquiterpene, (+)-epi-isozizaene (2), whose structure and stereochem. were detd. by a combination of 1H, 13C, COSY, HMQC, HMBC, and NOESY NMR. The steady-state kinetic parameters were kcat 0.049±0.001 s-1 and a Km (FPP) of 147±14 nM. Individual incubations of recombinant epi-isozizaene synthase with [1,1-2H2]FPP (1a), (1R)-[1-2H]-FPP (1b), and (1S)-[1-2H]-FPP (1c) and NMR anal. of the resulting deuterated epi-isozizaenes supported an isomerization-cyclization-rearrangement mechanism involving the intermediacy of (3R)-nerolidyl diphosphate (3).
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    36. 36
      Corre, C.; Song, L.; O’Rourke, S.; Chater, K. F.; Challis, G. L. 2-Alkyl-4-hydroxymethylfuran-3-carboxylic acids, antibiotic production inducers discovered by Streptomyces coelicolor genome mining. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 17510– 17515,  DOI: 10.1073/pnas.0805530105
      36
      2-Alkyl-4-hydroxymethylfuran-3-carboxylic acids, antibiotic production inducers discovered by Streptomyces coelicolor genome mining
      Corre, Christophe; Song, Lijiang; O'Rourke, Sean; Chater, Keith F.; Challis, Gregory L.
      Proceedings of the National Academy of Sciences of the United States of America (2008), 105 (45), 17510-17515CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)
      All of the genetic elements necessary for the prodn. of the antibiotic methylenomycin (Mm) and its regulation are contained within the 22-kb mmy-mmf gene cluster, which is located on the 356-kb linear plasmid SCP1 of Streptomyces coelicolor A3(2). A putative operon of 3 genes within this gene cluster, mmfLHP, was proposed to direct the biosynthesis of an A-factor-like signaling mol., which could play a role in the regulation of Mm biosynthesis. The mmfLHP operon was expressed under the control of its native promoter in S. coelicolor M512, a host lacking the SCP1 plasmid, and the ability to produce prodiginine and actinorhodin antibiotics. Comparative metabolic profiling led to the identification and structure elucidation of a family of 5 new 2-alkyl-4-hydroxymethylfuran-3-carboxylic acids (AHFCAs), collectively termed Mm furans (MMFs), as the products of the mmfLHP genes. MMFs specifically induce the prodn. of the Mm antibiotics in S. coelicolor. Comparative genomics analyses and searches of the natural product chem. literature indicated that other streptomycetes may produce AHFCAs, suggesting that they could form a general class of antibiotic biosynthesis inducers in Streptomyces species, with analogous functions to the better known γ-butyrolactone regulatory mols.
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    37. 37
      Wu, C.; Ichinose, K.; Choi, Y. H.; van Wezel, G. P. Aromatic polyketide GTRI-02 is a previously unidentified product of the act gene cluster in Streptomyces coelicolor A3(2). ChemBioChem 2017, 18, 1428– 1434,  DOI: 10.1002/cbic.201700107
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      Aromatic Polyketide GTRI-02 is a Previously Unidentified Product of the act Gene Cluster in Streptomyces coelicolor A3(2)
      Wu, Changsheng; Ichinose, Koji; Choi, Young Hae; van Wezel, Gilles P.
      ChemBioChem (2017), 18 (14), 1428-1434CODEN: CBCHFX; ISSN:1439-4227. (Wiley-VCH Verlag GmbH & Co. KGaA)
      The biosynthesis of arom. polyketides derived from type II polyketide synthases (PKSs) is complex, and it is not uncommon that highly similar gene clusters give rise to diverse structural architectures. The act biosynthetic gene cluster (BGC) of the model actinomycete Streptomyces coelicolor A3(2) is an archetypal type II PKS. Here we show that the act BGC also specifies the arom. polyketide GTRI-02 (1) and propose a mechanism for the biogenesis of its 3,4-dihydronaphthalen-1(2H)-one backbone. Polyketide 1 was also produced by Streptomyces sp. MBT76 after activation of the act-like qin gene cluster by overexpression of the pathway-specific activator. Mining of this strain also identified dehydroxy-GTRI-02 (2), which most likely originated from dehydration of 1 during the isolation process. This work shows that even extensively studied model gene clusters such as act of S. coelicolor can still produce new chem., offering new perspectives for drug discovery.
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    38. 38
      Li, B.; Cooper, L. E.; van der Donk, W. A. In Vitro Studies of Lantibiotic Biosynthesis. In Methods Enzymol.; Elsevier, 2009; Vol. 458, pp 533– 558.
      There is no corresponding record for this reference.
    39. 39
      Küsters, E.; Allgaier, H.; Jung, G.; Bayer, E. Resolution of sulphur-containing amino acids by chiral phase gas chromatography. Chromatographia 1984, 18, 287– 293,  DOI: 10.1007/BF02259079
      39
      Resolution of sulfur-containing amino acids by chiral phase gas chromatography
      Kuesters, E.; Allgaier, H.; Jung, G.; Bayer, E.
      Chromatographia (1984), 18 (6), 287-93CODEN: CHRGB7; ISSN:0009-5893.
      Me esters of the pentafluoropropionyl derivs. of lanthionines, cystathionines, and β-methyllanthionines were resolved by gas chromatog. on glass capillaries coated with the chiral stationary phase N-propionyl-L-valine tert-butylamide-polysiloxane (Chirasil-Val) within 35 min. L-Cystathionine elutes before its D-enantiomer in contrast to the usual order of emergence on an L-phase. The method was applied to the polypeptide antibiotic nisin, which contains meso-lanthionine and (2S,3S,6R)-3-methyllanthionine. N-Pentafluoropropionyl-S-alkylthiocysteine Me esters (alkyl = Me, Et, n- and iso-Pr, n- and sec-Bu, n-octyl, neo-pentyl, cyclohexyl, benzyl, tolyl) were sepd. on Chirasil-Val within 30 min. The identity of all derivs. was shown by combined gas chromatog.-mass spectrometry.
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    40. 40
      Liu, W.; Chan, A. S. H.; Liu, H.; Cochrane, S. A.; Vederas, J. C. Solid supported chemical syntheses of both components of the lantibiotic lacticin 3147. J. Am. Chem. Soc. 2011, 133, 14216– 14219,  DOI: 10.1021/ja206017p
      40
      Solid supported chemical syntheses of both components of the lantibiotic lacticin 3147
      Liu, Wei; Chan, Alice S. H.; Liu, Hongqiang; Cochrane, Stephen A.; Vederas, John C.
      Journal of the American Chemical Society (2011), 133 (36), 14216-14219CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lantibiotics are antimicrobial peptides produced by bacteria. Some are employed for food preservation, whereas others have therapeutic potential due to their activity against organisms resistant to current antibiotics. They are ribosomally synthesized and posttranslationally modified by dehydration of serine and threonine residues followed by attack of thiols of cysteines to form monosulfide lanthionine and methyllanthionine rings, resp. Chem. synthesis of peptide analogs is a powerful method to verify stereochem. and access structure-activity relationships. However, solid supported synthesis of lantibiotics has been difficult due to problems in generating lanthionines and methyllanthionines with orthogonal protection and good stereochem. control. We report the solid-phase syntheses of both peptides of a two-component lantibiotic, lacticin 3147. Both successive and interlocking ring systems were synthesized on-resin, thereby providing a general methodol. for this family of natural products.
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    41. 41
      Zallot, R.; Oberg, N.; Gerlt, J. A. The EFI web resource for genomic enzymology tools: Leveraging protein, genome, and metagenome databases to discover novel enzymes and metabolic pathways. Biochemistry 2019, 58, 4169– 4182,  DOI: 10.1021/acs.biochem.9b00735
      41
      The EFI Web Resource for Genomic Enzymology Tools: Leveraging Protein, Genome, and Metagenome Databases to Discover Novel Enzymes and Metabolic Pathways
      Zallot, Remi; Oberg, Nils; Gerlt, John A.
      Biochemistry (2019), 58 (41), 4169-4182CODEN: BICHAW; ISSN:0006-2960. (American Chemical Society)
      The assignment of functions to uncharacterized proteins discovered in genome projects requires easily accessible tools and computational resources for large-scale, user-friendly leveraging of the protein, genome, and metagenome databases by experimentalists. This article describes the web resource developed by the Enzyme Function Initiative (EFI; accessed at https://efi.igb.illinois.edu/) that provides "genomic enzymol." tools ("web tools") for (1) generating sequence similarity networks (SSNs) for protein families (EFI-EST); (2) analyzing and visualizing genome context of the proteins in clusters in SSNs (in genome neighborhood networks, GNNs, and genome neighborhood diagrams, GNDs) (EFI-GNT); and (3) prioritizing uncharacterized SSN clusters for functional assignment based on metagenome abundance (chem. guided functional profiling, CGFP) (EFI-CGFP). The SSNs generated by EFI-EST are used as the input for EFI-GNT and EFI-CGFP, enabling easy transfer of information among the tools. The networks are visualized and analyzed using Cytoscape, a widely used desktop application; GNDs and CGFP heatmaps summarizing metagenome abundance are viewed within the tools. We provide a detailed example of the integrated use of the tools with an anal. of glycyl radical enzyme superfamily (IPR004184) found in the human gut microbiome. This anal. demonstrates that (1) SwissProt annotations are not always correct, (2) large-scale genome context analyses allow the prediction of novel metabolic pathways, and (3) metagenome abundance can be used to identify/prioritize uncharacterized proteins for functional investigation.
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    42. 42
      Gerlt, J. A.; Bouvier, J. T.; Davidson, D. B.; Imker, H. J.; Sadkhin, B.; Slater, D. R.; Whalen, K. L. Enzyme Function Initiative-Enzyme Similarity Tool (EFI-EST): A web tool for generating protein sequence similarity networks. Biochim. Biophys. Acta, Proteins Proteomics 2015, 1854, 1019– 1037,  DOI: 10.1016/j.bbapap.2015.04.015
      42
      Enzyme function initiative-enzyme similarity tool (EFI-EST): A web tool for generating protein sequence similarity networks
      Gerlt, John A.; Bouvier, Jason T.; Davidson, Daniel B.; Imker, Heidi J.; Sadkhin, Boris; Slater, David R.; Whalen, Katie L.
      Biochimica et Biophysica Acta, Proteins and Proteomics (2015), 1854 (8), 1019-1037CODEN: BBAPBW; ISSN:1570-9639. (Elsevier B. V.)
      A review. The Enzyme Function Initiative, an NIH/NIGMS-supported Large-Scale Collaborative Project (EFI; U54GM093342; http://enzymefunction.org/), is focused on devising and disseminating bioinformatics and computational tools as well as exptl. strategies for the prediction and assignment of functions (in vitro activities and in vivo physiol./metabolic roles) to uncharacterized enzymes discovered in genome projects. Protein sequence similarity networks (SSNs) are visually powerful tools for analyzing sequence relationships in protein families (H.J. Atkinson, J.H. Morris, T.E. Ferrin, and P.C. Babbitt, PLoS One 2009, 4, e4345). However, the members of the biol./biomedical community have not had access to the capability to generate SSNs for their "favorite" protein families. In this article we announce the EFI-EST (Enzyme Function Initiative-Enzyme Similarity Tool) web tool (http://efi.igb.illinois.edu/efi-est/) that is available without cost for the automated generation of SSNs by the community. The tool can create SSNs for the "closest neighbors" of a user-supplied protein sequence from the UniProt database (Option A) or of members of any user-supplied Pfam and/or InterPro family (Option B). We provide an introduction to SSNs, a description of EFI-EST, and a demonstration of the use of EFI-EST to explore sequence-function space in the OMP decarboxylase superfamily (PF00215). This article is designed as a tutorial that will allow members of the community to use the EFI-EST web tool for exploring sequence/function space in protein families.
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    43. 43
      Le, T.; Fouque, K. J. D.; Santos-Fernandez, M.; Navo, C. D.; Jiménez-Osés, G.; Sarksian, R.; Fernandez-Lima, F. A.; van Der Donk, W. A. Substrate sequence controls regioselectivity of lanthionine formation by ProcM. J. Am. Chem. Soc. 2021, 143, 18733– 18743,  DOI: 10.1021/jacs.1c09370
      43
      Substrate Sequence Controls Regioselectivity of Lanthionine Formation by ProcM
      Le, Tung; Jeanne Dit Fouque, Kevin; Santos-Fernandez, Miguel; Navo, Claudio D.; Jimenez-Oses, Gonzalo; Sarksian, Raymond; Fernandez-Lima, Francisco Alberto; van der Donk, Wilfred A.
      Journal of the American Chemical Society (2021), 143 (44), 18733-18743CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)
      Lanthipeptides belong to the family of ribosomally synthesized and post-translationally modified peptides (RiPPs). The (methyl)lanthionine crosslinks characteristic to lanthipeptides are essential for their stability and bioactivities. In most bacteria, lanthipeptides are maturated from single precursor peptides encoded in the corresponding biosynthetic gene clusters. However, cyanobacteria engage in combinatorial biosynthesis and encode as many as 80 substrate peptides with highly diverse sequences that are modified by a single lanthionine synthetase into lanthipeptides of different lengths and ring patterns. It is puzzling how a single enzyme could exert control over the cyclization processes of such a wide range of substrates. Here, the authors used a library of ProcA3.3 precursor peptide variants and show that it is not the enzyme ProcM but rather its substrate sequences that det. the regioselectivity of lanthionine formation. The authors also demonstrate the utility of trapped ion mobility spectrometry-tandem mass spectrometry (TIMS-MS/MS) as a fast and convenient method to efficiently sep. lanthipeptide constitutional isomers, particularly in cases where the isomers cannot be resolved by conventional liq. chromatog. The authors' data allowed identification of factors that are important for the cyclization outcome, but also showed that there are no easily identifiable predictive rules for all sequences. The authors' findings provide a platform for future deep learning approaches to allow such prediction of ring patterns of products of combinatorial biosynthesis.
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    44. 44
      Bobeica, S. C.; Zhu, L.; Acedo, J. Z.; Tang, W.; van der Donk, W. A. Structural determinants of macrocyclization in substrate-controlled lanthipeptide biosynthetic pathways. Chem. Sci. 2020, 11, 12854– 12870,  DOI: 10.1039/D0SC01651A
      44
      Structural determinants of macrocyclization in substrate-controlled lanthipeptide biosynthetic pathways
      Bobeica, Silvia C.; Zhu, Lingyang; Acedo, Jeella Z.; Tang, Weixin; van der Donk, Wilfred A.
      Chemical Science (2020), 11 (47), 12854-12870CODEN: CSHCCN; ISSN:2041-6520. (Royal Society of Chemistry)
      Lanthipeptides are characterized by thioether crosslinks formed by post-translational modifications. The cyclization process that favors a single ring pattern over many other possible ring patterns has been the topic of much speculation. Recent studies suggest that for some systems the cyclization pattern and stereochem. is detd. not by the enzyme, but by the sequence of the precursor peptide. However, the factors that govern the outcome of the cyclization process are not understood. This study presents the three-dimensional structures of seven lanthipeptides detd. by NMR spectroscopy, including five prochlorosins and the two peptides that make up cytolysin, a virulence factor produced by Enterococcus faecalis that is directly linked to human disease. These peptides were chosen because their substrate sequence dets. either the ring pattern (prochlorosins) or the stereochem. of cyclization (cytolysins). We present the structures of prochlorosins 1.1, 2.1, 2.8, 2.10 and 2.11, the first three-dimensional structures of prochlorosins. Our findings provide insights into the mol. determinants of cyclization as well as why some prochlorosins may be better starting points for library generation than others. The structures of the large and small subunits of the enterococcal cytolysin show that these peptides have long helical stretches, a rare observation for lanthipeptides characterized to date. These helixes may explain their pore forming activity and suggest that the small subunit may recognize a mol. target followed by recruitment of the large subunit to span the membrane.
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    45. 45
      Altschul, S. F.; Madden, T. L.; Schaffer, A. A.; Zhang, J.; Zhang, Z.; Miller, W.; Lipman, D. J. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997, 25, 3389– 3402,  DOI: 10.1093/nar/25.17.3389
      45
      Gapped BLAST and PSI-BLAST: a new generation of protein database search programs
      Altschul, Stephen F.; Madden, Thomas L.; Schaffer, Alejandro A.; Zhang, Jinghui; Zhang, Zheng; Miller, Webb; Lipman, David J.
      Nucleic Acids Research (1997), 25 (17), 3389-3402CODEN: NARHAD; ISSN:0305-1048. (Oxford University Press)
      The BLAST programs are widely used tools for searching protein and DNA databases for sequence similarities. For protein comparisons, a variety of definitional, algorithmic and statistical refinements described here permits the execution time of the BLAST programs to be decreased substantially while enhancing their sensitivity to weak similarities. A new criterion for triggering the extension of word hits, combined with a new heuristic for generating gapped alignments, yields a gapped BLAST program that runs at approx. three times the speed of the original. In addn., a method is introduced for automatically combining statistically significant alignments produced by BLAST into a position-specific score matrix, and searching the database using this matrix. The resulting Position-Specific Iterated BLAST (PSI-BLAST) program runs at approx. the same speed per iteration as gapped BLAST, but in many cases is much more sensitive to weak but biol. relevant sequence similarities. PSI-BLAST is used to uncover several new and interesting members of the BRCT superfamily. The source code for the new BLAST programs is available by anonymous ftp from the machine ncbi.nlm.nih.gov, within the directory 'blast', and the programs may be run from NCBIs web site at http://www.ncbi.nlm.nih.gov/.
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  • Supporting Information

    Supporting Information

    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acschembio.2c00492.

    • Glutamyl lyase SSN (ZIP)

    • Experimental procedures; extended experimental data and bioinformatic analysis; materials and methods; primers used in this study; observed and calculated m/z ratios for fragments of GluC-digested mCoiA1; NEM alkylation assay for CoiA1 co-expressed with CoiB-H994A, CoiC, and CoiSA(ED); LC-MS analysis of the GluC digestion product of mCoiA1 and variants; description table of orthologs obtained from PSI-BLAST of OlvA (PDF)

    Accession Codes

    NCBI protein accessions: CoiA1, WP_011031313.1; CoiB, WP_011031312.1; CoiC, WP_011031311.1; and CoiSA, WP_011031310.1.


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