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Production of Sactipeptides in Escherichia coli: Probing the Substrate Promiscuity of Subtilosin A Biosynthesis

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Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy, Chapel Hill, North Carolina, United States
Cite this: ACS Chem. Biol. 2016, 11, 6, 1737–1744
Publication Date (Web):March 28, 2016
Copyright © 2016 American Chemical Society

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    Sactipeptides are peptide-derived natural products that are processed by remarkable, radical-mediated cysteine sulfur to α-carbon coupling reactions. The resulting sactionine thioether linkages give rise to the unique defined structures and concomitant biological activities of sactipeptides. An E. coli heterologous expression system, based on the biosynthesis of one such sactipeptide, subtilosin A, is described and this expression system is exploited to probe the promiscuity of the subtilosin A sactionine bond-forming enzyme, AlbA. These efforts allowed the facile expression and isolation of a small library of mutant sactipeptides based on the subtilosin A precursor peptide, demonstrating broad substrate promiscuity where none was previously known. Importantly, we show that the positions of the sactionine linkages can be moved, giving rise to new, unnatural sactipeptide structures. E. coli heterologous expression also allowed incorporation of unnatural amino acids into sactipeptides by means of amber-suppression technology, potentially opening up new chemistry and new applications for unnatural sactipeptides.

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    8. Tyler L. Grove, Paul M. Himes, Sungwon Hwang, Hayretin Yumerefendi, Jeffrey B. Bonanno, Brian Kuhlman, Steven C. Almo, and Albert A. Bowers . Structural Insights into Thioether Bond Formation in the Biosynthesis of Sactipeptides. Journal of the American Chemical Society 2017, 139 (34) , 11734-11744.
    9. Brandon J. Burkhart, Nidhi Kakkar, Graham A. Hudson, Wilfred A. van der Donk, and Douglas A. Mitchell . Chimeric Leader Peptides for the Generation of Non-Natural Hybrid RiPP Products. ACS Central Science 2017, 3 (6) , 629-638.
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    12. He Li, Wei Ding, Qi Zhang. Discovery and engineering of ribosomally synthesized and post-translationally modified peptide (RiPP) natural products. RSC Chemical Biology 2024, 5 (2) , 90-108.
    13. . Peptide Natural Products I: RiPPs. 2022, 100-149.
    14. Kenzie A. Clark, Brett C. Covington, Mohammad R. Seyedsayamdost. Biosynthesis-guided discovery reveals enteropeptins as alternative sactipeptides containing N-methylornithine. Nature Chemistry 2022, 14 (12) , 1390-1398.
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    20. A. Alloul, W. Van Kampen, M. Cerruti, S. Wittouck, M. Pabst, D.G. Weissbrodt. Exploring the role of antimicrobials in the selective growth of purple phototrophic bacteria through genome mining and agar spot assays. Letters in Applied Microbiology 2022, 75 (5) , 1275-1285.
    21. Emerson Glassey, Andrew M. King, Daniel A. Anderson, Zhengan Zhang, Christopher A. Voigt, . Functional expression of diverse post-translational peptide-modifying enzymes in Escherichia coli under uniform expression and purification conditions. PLOS ONE 2022, 17 (9) , e0266488.
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    23. Sebastian L. Wenski, Sirinthra Thiengmag, Eric J.N. Helfrich. Complex peptide natural products: Biosynthetic principles, challenges and opportunities for pathway engineering. Synthetic and Systems Biotechnology 2022, 7 (1) , 631-647.
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    40. Graham A Hudson, Douglas A Mitchell. RiPP antibiotics: biosynthesis and engineering potential. Current Opinion in Microbiology 2018, 45 , 61-69.
    41. Yi Zhang, Manyun Chen, Steven D. Bruner, Yousong Ding. Heterologous Production of Microbial Ribosomally Synthesized and Post-translationally Modified Peptides. Frontiers in Microbiology 2018, 9
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