ACS Publications. Most Trusted. Most Cited. Most Read
My Activity

Glycosylation Is Important for Binding to Human Calcitonin Receptors

View Author Information
Division of Molecular Medicine, Department of Medicine, Cornell University Medical College and The New York Hospital, New York, New York 10021
Cite this: Biochemistry 1999, 38, 6, 1866–1872
Publication Date (Web):January 21, 1999
Copyright © 1999 American Chemical Society

    Article Views





    Other access options


    Human calcitonin receptor (hCTR) subtypes contain three or four potential Asn-linked glycosylation sites in their extracellular amino termini. The role of glycosylation in hCTR function has not been identified, but it has been suggested that inhibition of glycosylation does not affect binding or signaling. To determine the role of glycosylation in hCTR biology, we studied the effects of inhibition of glycosylation and of substitution of Asn residues that are potential glycosylation sites. Native and mutated hCTRs were studied after transient expression in monkey kidney COS-1 cells. Tunicamycin, administered as part of a treatment protocol that inhibited glycosylation of all expressed receptors, decreased salmon calcitonin (sCT) binding affinities and signaling potencies at hCTRs with three or four potential glycosylation sites. In hCTR3, which contains three potential glycosylation sites at positions 26, 78, and 83, site-specific substitution of Asn-26 by Ala had no effect on sCT binding affinity or potency, whereas substitution of Asn-78 or Asn-83 lowered sCT affinity and potency. A mutant hCTR3 in which all three Asn residues were substituted with Ala exhibited no high-affinity sCT binding and potencies of several calcitonin analogues that were more than 100-fold lower than that of native hCTR3. Our data show that glycosylation is important for high-affinity binding and potency of calcitonin analogues at hCTRs.

    Read this article

    To access this article, please review the available access options below.

    Get instant access

    Purchase Access

    Read this article for 48 hours. Check out below using your ACS ID or as a guest.


    Access through Your Institution

    You may have access to this article through your institution.

    Your institution does not have access to this content. You can change your affiliated institution below.

     This work was supported by U.S. Public Health Service Grants DK43036 (to M.C.G.) and DK50673 (to D.R.N.).


     To whom correspondence should be addressed:  Cornell University Medical College, 1300 York Ave., Room A328, New York, NY 10021. Telephone:  (212) 746-6275. Fax:  (212) 746-6289. E-mail:  mcgersh@

    Cited By

    This article is cited by 40 publications.

    1. Sang-Min Lee, Jason M. Booe, Joseph J. Gingell, Virginie Sjoelund, Debbie L. Hay, and Augen A. Pioszak . N-Glycosylation of Asparagine 130 in the Extracellular Domain of the Human Calcitonin Receptor Significantly Increases Peptide Hormone Affinity. Biochemistry 2017, 56 (26) , 3380-3393.
    2. Y. Yan, D. J. Scott, T. N. Wilkinson, J. Ji, G. W. Tregear and R. A. D. Bathgate . Identification of the N-Linked Glycosylation Sites of the Human Relaxin Receptor and Effect of Glycosylation on Receptor Function. Biochemistry 2008, 47 (26) , 6953-6968.
    3. Jana Klose,, Klaus Fechner,, Michael Beyermann,, Eberhard Krause,, Norbert Wendt,, Michael Bienert,, Rainer Rudolph, and, Sven Rothemund. Impact of N-Terminal Domains for Corticotropin-Releasing Factor (CRF) Receptor−Ligand Interactions. Biochemistry 2005, 44 (5) , 1614-1623.
    4. Jian Ma, Liang Yan, Jingkui Yang, Yujian He, Li Wu. Effect of Modification Strategies on the Biological Activity of Peptides/Proteins. ChemBioChem 2024, 25 (3)
    5. Silja Mordhorst, Fleur Ruijne, Anna L. Vagstad, Oscar P. Kuipers, Jörn Piel. Emulating nonribosomal peptides with ribosomal biosynthetic strategies. RSC Chemical Biology 2023, 4 (1) , 7-36.
    6. Sepideh Kalhor, Alireza Fattahi. Design of amino acid- and carbohydrate-based anticancer drugs to inhibit polymerase η. Scientific Reports 2022, 12 (1)
    7. Camille Vaubourg, Evelyne Gicquel, Isabelle Richard, William Lostal, Jessica Bellec, , . Minimal Consequences of CMAH and DBA/2 Backgrounds on a FKRP Deficient Model. Journal of Neuromuscular Diseases 2021, 8 (5) , 785-793.
    8. Weiwei Wang, Yanan Tian, Xiaoliu Shi, Qiang Ma, Yue Xu, Gangjie Yang, Wen Yi, Ying Shi, Naiming Zhou. N‐glycosylation of the human neuropeptide QRFP receptor (QRFPR) is essential for ligand binding and receptor activation. Journal of Neurochemistry 2021, 158 (2) , 138-152.
    9. Sang-Min Lee, Yejin Jeong, John Simms, Margaret L. Warner, David R. Poyner, Ka Young Chung, Augen A. Pioszak. Calcitonin Receptor N-Glycosylation Enhances Peptide Hormone Affinity by Controlling Receptor Dynamics. Journal of Molecular Biology 2020, 432 (7) , 1996-2014.
    10. Parvaiz Ahmad Dar, Usma Manzoor, Snowber Shabir Wani, Fasil Ali, Tanveer Ali Dar. Role of Glycosylation in Modulating Therapeutic Efficiency of Protein Pharmaceuticals. 2019, 131-143.
    11. Tyler J. Wadzinski, Angela Steinauer, Liana Hie, Guillaume Pelletier, Alanna Schepartz, Scott J. Miller. Rapid phenolic O-glycosylation of small molecules and complex unprotected peptides in aqueous solvent. Nature Chemistry 2018, 10 (6) , 644-652.
    12. John Simms, Sarah Routledge, Romez Uddin, David Poyner. The Structure of the CGRP and Related Receptors. 2018, 23-36.
    13. Yi-Lynn Liang, Maryam Khoshouei, Mazdak Radjainia, Yan Zhang, Alisa Glukhova, Jeffrey Tarrasch, David M. Thal, Sebastian G. B. Furness, George Christopoulos, Thomas Coudrat, Radostin Danev, Wolfgang Baumeister, Laurence J. Miller, Arthur Christopoulos, Brian K. Kobilka, Denise Wootten, Georgios Skiniotis, Patrick M. Sexton. Phase-plate cryo-EM structure of a class B GPCR–G-protein complex. Nature 2017, 546 (7656) , 118-123.
    14. Sang-Min Lee, Debbie L. Hay, Augen A. Pioszak. Calcitonin and Amylin Receptor Peptide Interaction Mechanisms. Journal of Biological Chemistry 2016, 291 (16) , 8686-8700.
    15. Shayli Varasteh Moradi, Waleed M. Hussein, Pegah Varamini, Pavla Simerska, Istvan Toth. Glycosylation, an effective synthetic strategy to improve the bioavailability of therapeutic peptides. Chemical Science 2016, 7 (4) , 2492-2500.
    16. T Qi, M Dong, HA Watkins, D Wootten, LJ Miller, DL Hay. Receptor activity‐modifying protein‐dependent impairment of calcitonin receptor splice variant Δ(1–47) hCT (a) function. British Journal of Pharmacology 2013, 168 (3) , 644-657.
    17. Nobuo Suzuki, Toshio Sekiguchi, Honoo Satake, Kanoko Kato, Yudai Nishiyama, Hideya Takahashi, Janine A. Danks, T. John Martin, Atsuhiko Hattori, Masaki Nakano, Makiko Kakikawa, Sotoshi Yamada, Maho Ogoshi, Susumu Hyodo, Yoko Yamaguchi, Vishwajit S. Chowdhury, Kazuichi Hayakawa, Hisayuki Funahashi, Tatsuya Sakamoto, Yuichi Sasayama. Cloning of two members of the calcitonin-family receptors from stingray, Dasyatis akajei: Possible physiological roles of the calcitonin family in osmoregulation. Gene 2012, 499 (2) , 326-331.
    18. Toni Segovia-Silvestre, Caroline Bonnefond, Bodil C Sondergaard, Tjorbjoern Christensen, Morten A Karsdal, Anne C Bay-Jensen. Identification of the calcitonin receptor in osteoarthritic chondrocytes. BMC Research Notes 2011, 4 (1)
    19. Quan Chen, Laurence J. Miller, Maoqing Dong. Role of N -linked glycosylation in biosynthesis, trafficking, and function of the human glucagon-like peptide 1 receptor. American Journal of Physiology-Endocrinology and Metabolism 2010, 299 (1) , E62-E68.
    20. Vi Pham, Patrick M. Sexton. Photoaffinity scanning in the mapping of the peptide receptor interface of class II G protein—coupled receptors. Journal of Peptide Science 2004, 10 (4) , 179-203.
    21. Vi Pham, John D. Wade, Patrick M. Sexton. Application of photoaffinity crosslinking in determining the interaction between calcitonin and its receptor. International Journal of Peptide Research and Therapeutics 2003, 10 (5-6) , 447-453.
    22. Vi Pham, John D. Wade, Patrick M. Sexton. Application of photoaffinity crosslinking in determining the interaction between calcitonin and its receptor. Letters in Peptide Science 2003, 10 (5-6) , 447-453.
    23. Angela M. Inzerillo, Mone Zaidi, Christopher L.-H. Huang. Calcitonin: The Other Thyroid Hormone. Thyroid 2002, 12 (9) , 791-798.
    24. M Zaidi, A.M Inzerillo, B.S Moonga, P.J.R Bevis, C.L.-H Huang. Forty years of calcitonin—where are we now? A tribute to the work of Iain Macintyre, FRS. Bone 2002, 30 (5) , 655-663.
    25. Walter Born, Roman Muff, Jan A. Fischer. Functional interaction of G protein‐coupled receptors of the adrenomedullin peptide family with accessory receptor‐activity‐modifying proteins (RAMP). Microscopy Research and Technique 2002, 57 (1) , 14-22.
    26. Deborah L. Galson, Steven R. Goldring. Structure and Molecular Biology of the Calcitonin Receptor. 2002, 603-617.
    27. Mone Zaidi, Angela M. Inzerillo, Bruce Troen, Baljit S. Moonga, Etsuko Abe, Peter Burckhardt. Molecular and Clinical Pharmacology of Calcitonin. 2002, 1423-1440.
    28. B. W. Purdue, N. Tilakaratne, P. M. Sexton. Molecular Pharmacology of the Calcitonin Receptor. Receptors and Channels 2002, 8 (3-4) , 243-255.
    29. W. Born, J. A. Fischer, R. Muff. Receptors for Calcitonin Gene-Related Peptide, Adrenomedullin, and Amylin: The Contributions of Novel Receptor-Activity-Modifying Proteins. Receptors and Channels 2002, 8 (3-4) , 201-209.
    30. Roman Muff, Walter Born, Jan A Fischer. Adrenomedullin and related peptides: receptors and accessory proteins. Peptides 2001, 22 (11) , 1765-1772.
    31. Iman Q. Assil, Abdul B. Abou-Samra. N-glycosylation of CRF receptor type 1 is important for its ligand-specific interaction. American Journal of Physiology-Endocrinology and Metabolism 2001, 281 (5) , E1015-E1021.
    32. Kathy Yuen-Yee Chan, Ronald Ting-Kai Pang, Billy Kwok-Chong Chow. Functional Segregation of the Highly Conserved Basic Motifs within the Third Endoloop of the Human Secretin Receptor. Endocrinology 2001, 142 (9) , 3926-3934.
    33. Yong-Jiang Cao, Gerald Gimpl. A constitutively active pituitary adenylate cyclase activating polypeptide (PACAP) type I receptor shows enhanced photoaffinity labeling of its highly glycosylated form. Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology 2001, 1548 (1) , 139-151.
    34. Shigeki Kamitani, Tsuneaki Sakata. Glycosylation of human CRLR at Asn123 is required for ligand binding and signaling. Biochimica et Biophysica Acta (BBA) - Molecular Cell Research 2001, 1539 (1-2) , 131-139.
    35. Xi-Qin Ding, Vesile Dolu, Elizabeth M. Hadac, Eileen L. Holicky, Delia I. Pinon, Terry P. Lybrand, Laurence J. Miller. Refinement of the Structure of the Ligand-occupied Cholecystokinin Receptor Using a Photolabile Amino-terminal Probe. Journal of Biological Chemistry 2001, 276 (6) , 4236-4244.
    36. Nicole Bühlmann, Amaya Aldecoa, Kerstin Leuthäuser, Remo Gujer, Roman Muff, Jan A Fischer, Walter Born. Glycosylation of the calcitonin receptor‐like receptor at Asn 60 or Asn 112 is important for cell surface expression. FEBS Letters 2000, 486 (3) , 320-324.
    37. Richard V. Benya, Takashi Kusui, Tatsuro Katsuno, Takaharu Tsuda, Samuel A. Mantey, James F. Battey, Robert T. Jensen. Glycosylation of the Gastrin-Releasing Peptide Receptor and Its Effect on Expression, G Protein Coupling, and Receptor Modulatory Processes. Molecular Pharmacology 2000, 58 (6) , 1490-1501.
    38. Nobuo Suzuki, Tohru Suzuki, Tadahide Kurokawa. Cloning of a calcitonin gene-related peptide receptor and a novel calcitonin receptor-like receptor from the gill of flounder, Paralichthys olivaceus. Gene 2000, 244 (1-2) , 81-88.
    39. Ronald Ting-Kai Pang, Samuel Sai-Ming Ng, Christopher Hon-Ki Cheng, Martin H. Holtmann, Laurence J. Miller, Billy Kwok-Chong Chow. Role of N-Linked Glycosylation on the Function and Expression of the Human Secretin Receptor. Endocrinology 1999, 140 (11) , 5102-5111.
    40. Hao H. Ho, Dongyi Du, Marvin C. Gershengorn. The N Terminus of Kaposi's Sarcoma-associated Herpesvirus G Protein-coupled Receptor Is Necessary for High Affinity Chemokine Binding but Not for Constitutive Activity. Journal of Biological Chemistry 1999, 274 (44) , 31327-31332.

    Pair your accounts.

    Export articles to Mendeley

    Get article recommendations from ACS based on references in your Mendeley library.

    Pair your accounts.

    Export articles to Mendeley

    Get article recommendations from ACS based on references in your Mendeley library.

    You’ve supercharged your research process with ACS and Mendeley!

    STEP 1:
    Click to create an ACS ID

    Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

    Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

    Please note: If you switch to a different device, you may be asked to login again with only your ACS ID.

    Your Mendeley pairing has expired. Please reconnect