title: Toward the active conformation of insulin: stereospecific modulation of a structural switch in the B chain.
abstract: How insulin binds to the insulin receptor has long been a subject of speculation. Although the structure of the free hormone has been extensively characterized, a variety of evidence suggests that a conformational change occurs upon receptor binding. Here, we employ chiral mutagenesis, comparison of corresponding d and l amino acid substitutions, to investigate a possible switch in the B-chain. To investigate the interrelation of structure, function, and stability, isomeric analogs have been synthesized in which an invariant glycine in a beta-turn (Gly(B8)) is replaced by d- or l-Ser. The d substitution enhances stability (DeltaDeltaG(u) 0.9 kcal/mol) but impairs receptor binding by 100-fold; by contrast, the l substitution markedly impairs stability (DeltaDeltaG(u) -3.0 kcal/mol) with only 2-fold reduction in receptor binding. Although the isomeric structures each retain a native-like overall fold, the l-Ser(B8) analog exhibits fewer helix-related and long range nuclear Overhauser effects than does the d-Ser(B8) analog or native monomer. Evidence for enhanced conformational fluctuations in the unstable analog is provided by its attenuated CD spectrum. The inverse relationship between stereospecific stabilization and receptor binding strongly suggests that the B7-B10 beta-turn changes conformation on receptor binding.
authors.affiliation: Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106, USA.
authors.forename: Qing-Xin
authors.initials: QX
authors.lastname: Hua
authors.forename: Satoe
authors.initials: S
authors.lastname: Nakagawa
authors.forename: Shi-Quan
authors.initials: SQ
authors.lastname: Hu
authors.forename: Wenhua
authors.initials: W
authors.lastname: Jia
authors.forename: Shuhua
authors.initials: S
authors.lastname: Wang
authors.forename: Michael A
authors.initials: MA
authors.lastname: Weiss
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D006801:Humans; D007328:Insulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D011972:Receptor, Insulin / Q000378:metabolism
chemical_list: D007328:Insulin; D011972:Receptor, Insulin