A new B-chain mutant of insulin: comparison with the insulin crystal structure and role of sulfonate groups in the B-chain structure.
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.
Dupradeau
F-Y
FY
GRBPD, UPRES 2629, Faculté de Pharmacie et de Médecine, Amiens, France.
Richard
T
T
Le Flem
G
G
Oulyadi
H
H
Prigent
Y
Y
Monti
J-P
JP
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002942:Circular Dichroism; D007328:Insulin / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013451:Sulfonic Acids / Q000737:chemistry*
D007328:Insulin; D013451:Sulfonic Acids