title: Equilibrium denaturation of insulin and proinsulin.
abstract: The guanidine hydrochloride induced equilibrium denaturation of insulin and proinsulin was studied by using near- and far-ultraviolet (UV) circular dichroism (CD). The denaturation transition of insulin is reversible, cooperative, symmetrical, and the same whether detected by near- or far-UV CD. These results are consistent with a two-state denaturation process without any appreciable equilibrium intermediates. Analysis of the insulin denaturation data yields a Gibbs free energy of unfolding of 4.5 +/- 0.5 kcal/mol. Denaturation of proinsulin detected by near-UV CD appears to be the same as for insulin, but if detected by far-UV CD appears different. The far-UV CD results demonstrate a multiphasic transition with the connecting peptide portion unfolding at lower concentrations of denaturant. Similar studies with the isolated C-peptide show that its conformation and susceptibility to denaturation are independent of the rest of the proinsulin molecule. After the proinsulin denaturation results were adjusted for the connecting peptide contribution, a denaturation transition identical with that of insulin was obtained. These results show that for proinsulin, the connecting peptide segment is not a random coil; it is an autonomous folding unit, and the portion corresponding to insulin is identical with insulin in terms of conformational stability.
authors: Department of Pharmaceutical Research, Eli Lilly & Co., Indianapolis, Indiana 46285.
authors: D N
authors: DN
authors: Brems
authors: P L
authors: PL
authors: Brown
authors: L A
authors: LA
authors: Heckenlaible
authors: B H
authors: BH
authors: Frank
mesh_terms: D002942:Circular Dichroism; D000431:Ethanol / Q000494:pharmacology; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism*; D011384:Proinsulin / Q000378:metabolism*; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D013816:Thermodynamics
chemical_list: D006146:Guanidines; D007328:Insulin; D000431:Ethanol; D011384:Proinsulin; D019791:Guanidine