A new B‐chain mutant of insulin: comparison with the insulin crystal structure and role of sulfonate groups in the B‐chain structure

Abstract

The solution structure of a new B-chain mutant of bovine insulin, in which the cysteines B7 and B19 are replaced by two serines, has been determined by circular dichroism, 2D-NMR and molecular modeling. This structure is compared with that of the oxidized B-chain of bovine insulin [Hawkins et al. (1995) Int. J. Peptide Protein Res.46, 424-433]. Circular dichroism spectroscopy showed in particular that a higher percentage of helical secondary structure for the B-chain mutant is estimated in trifluoroethanol solution in comparison with the oxidized B-chain. 2D-NMR experiments confirmed, among multiple conformations, that the B-chain mutant presents defined secondary structures such as a alpha-helix between residues B9 and B19, and a beta-turn between amino acids B20 and B23 in aqueous trifluoroethanol. The 3D structures, which are consistent with NMR data and were obtained using a simulated annealing protocol, showed that the tertiary structure of the B-chain mutant is better resolved and is more in agreement with the insulin crystal structure than the oxidized B-chain structure described by Hawkins et al. An explanation could be the presence of two sulfonate groups in the oxidized insulin B-chain. Either by their charges and/or their size, such chemical groups could play a destructuring effect and thus could favor peptide flexibility and conformational averaging. Thus, this study provides new insights on the folding of isolated B-chains.

Intermolecular interactions between the neurotensin and the third extracellular loop of human neurotensin 1 receptor

“…Its sequence is as follows: Val 321 -Arg 322 -Arg 323 -Leu 324 -Met 325 -Phe 326 -Ser 327 -Tyr 328 -Ile 329 -Ser 330 -Asp 331 -Glu 332 -Gln 333 -Trp 334 -Thr 335 -Pro 336 -Phe 337 -Leu 338 -Tyr 339 -Asp 340 -Phe 341 -Tyr 342 -His 343 -Tyr 344 . To avoid thiol group instability in solution, we have replaced the Cys 327 by the residue serine (Dupradeau et al, 2002). It was synthesized and purified (purity $ 98%) by polypeptide (Strasbourg, France) and SynVec (Bordeaux, France).…”

Exploring the Implications of Vitamin B12 Conjugation to Insulin on Insulin Receptor Binding

“…[6] The a-helical geometry of the A chain (A1-A10 and A12-A20; A-Cys 11 is the non-helical residue involved in the intra-helical disulfide linkage to A-Cys 6) and central B chain segment (B9-B19) are preserved (as determined by NMR [7][8][9] and MD [10][11][12][13][14] studies) in all physiologically relevant conformations, while the remainder of the B chain (B1-B8 and B20-B30) is marked by significant structural differences between the dimeric/hexameric crystal and various solution-phase forms.…”

Human insulin A‐chain peptide analog(s) with in vitro biological activity

“…Therefore, several chemical groups such as S-thiomethyl, S-sulfonate, sulfonate, and hydroxyl have often been used to mimic the thiol group in studies of 3D structures of insulin isolated chains. [12][13][14] From these studies, it has been recently deduced that the negative charge of the sulfonate groups exerts a destructuring effect, destabilizing the 3D structure, and affecting the folding of an isolated insulin B-chain analog. In contrast, the serine hydroxyl group promotes a greater ''insulin-like'' conformation.…”