title: Diabetes-associated mutations in insulin: consecutive residues in the B chain contact distinct domains of the insulin receptor.
abstract: How insulin binds to and activates the insulin receptor has long been the subject of speculation. Of particular interest are invariant phenylalanine residues at consecutive positions in the B chain (residues B24 and B25). Sites of mutation causing diabetes mellitus, these residues occupy opposite structural environments: Phe(B25) projects from the surface of insulin, whereas Phe(B24) packs against the core. Despite these differences, site-specific cross-linking suggests that each contacts the insulin receptor. Photoactivatable derivatives of insulin containing respective p-azidophenylalanine substitutions at positions B24 and B25 were synthesized in an engineered monomer (DKP-insulin). On ultraviolet irradiation each derivative cross-links efficiently to the receptor. Packing of Phe(B24) at the receptor interface (rather than against the core of the hormone) may require a conformational change in the B chain. Sites of cross-linking in the receptor were mapped to domains by Western blot. Remarkably, whereas B25 cross-links to the C-terminal domain of the alpha subunit in accord with previous studies (Kurose, T., et al. (1994) J. Biol. Chem. 269, 29190-29197), the probe at B24 cross-links to its N-terminal domain (the L1 beta-helix). Our results demonstrate that consecutive residues in insulin contact widely separated sequences in the receptor and in turn suggest a revised interpretation of electron-microscopic images of the complex. By tethering the N- and C-terminal domains of the extracellular alpha subunit, insulin is proposed to stabilize an active conformation of the disulfide-linked transmembrane tyrosine kinase.
authors.affiliation: Department of Biochemistry, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4935, USA.
authors.forename: Bin
authors.initials: B
authors.lastname: Xu
authors.forename: Shi-Quan
authors.initials: SQ
authors.lastname: Hu
authors.forename: Ying-Chi
authors.initials: YC
authors.lastname: Chu
authors.forename: Kun
authors.initials: K
authors.lastname: Huang
authors.forename: Satoe H
authors.initials: SH
authors.lastname: Nakagawa
authors.forename: Jonathan
authors.initials: J
authors.lastname: Whittaker
authors.forename: Panayotis G
authors.initials: PG
authors.lastname: Katsoyannis
authors.forename: Michael A
authors.initials: MA
authors.lastname: Weiss
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001386:Azides / Q000737:chemistry; D015153:Blotting, Western; D016466:CHO Cells; D002918:Chymotrypsin / Q000737:chemistry; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003920:Diabetes Mellitus / Q000235:genetics*; D019281:Dimerization; D004220:Disulfides; D005091:Exons; D006801:Humans; D007328:Insulin / Q000235:genetics* / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010649:Phenylalanine / Q000031:analogs & derivatives / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011972:Receptor, Insulin / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D014357:Trypsin / Q000737:chemistry; D014466:Ultraviolet Rays
chemical_list: D001386:Azides; D003432:Cross-Linking Reagents; D004220:Disulfides; D007328:Insulin; C027162:4-azidophenylalanine; D010649:Phenylalanine; D011972:Receptor, Insulin; D002918:Chymotrypsin; D014357:Trypsin