title: Characterization of the N-methyltransferase activities of the multifunctional polypeptide cyclosporin synthetase.
abstract: This study demonstrates a critical role for N-methylation in cyclosporin biosynthesis and maintenance of the biologically active cyclosporin conformation. The structural requirements for the AdoMet binding to CySyn were defined. N-methylation of specific amide positions in the cyclosporin backbone is critical for the complete assembly and cyclization of the cyclosporin peptide. A maximum of two desmethyl positions is tolerated before peptide assembly stalls. Subinhibitory concentrations of AdoMet analogs directed peptide assembly towards cyclosporins with less than seven N-methylated amide bonds. Molecular modeling and nuclear magnetic resonance analyses indicate that N-methylation of specific amide bond positions in the cyclosporin backbone is mandatory for the formation of a product-like conformation and recognition by the acceptor site of the downstream peptide bond forming C-domain.
authors.affiliation: Medicinal Chemistry and Drug Action, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Australia. Tony.Velkov@monash.edu.au
authors.forename: Tony
authors.initials: T
authors.lastname: Velkov
authors.forename: James
authors.initials: J
authors.lastname: Horne
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Scanlon
authors.forename: Ben
authors.initials: B
authors.lastname: Capuano
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Yuriev
authors.forename: Alfons
authors.initials: A
authors.lastname: Lawen
mesh_terms: D000577:Amides / Q000378:metabolism; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism; D003067:Coenzymes / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D003500:Cyclization; D016572:Cyclosporine / Q000737:chemistry / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006860:Hydrogen Bonding; D008745:Methylation; D008780:Methyltransferases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry / Q000378:metabolism*; D010453:Peptide Synthases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012436:S-Adenosylmethionine / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D000577:Amides; D002352:Carrier Proteins; D003067:Coenzymes; D004791:Enzyme Inhibitors; D009097:Multienzyme Complexes; C048525:cyclosporin A synthetase; D012436:S-Adenosylmethionine; D016572:Cyclosporine; D008780:Methyltransferases; D010453:Peptide Synthases