Trifluoroethanol-induced conformational change of tetrameric and monomeric soybean agglutinin: Role of structural organization and implication for protein folding and stability

摘要

2,2,2-Trifuoroethanol (TFE)-induced conformational structure change of a beta-sheet legume lectin, soybean agglutinin (SBA) has been investigated employing its exclusive structural forms in quaternary (tetramer) and tertiary (monomer) states, by far- and near-UV CD, FTIR, fluorescence, low temperature phosphorescence and chemical modification. Far-UV CD results show that, for SBA tetramer, native atypical beta-conformation transforms to a highly alpha-helical structure, with the helical content reaching 57 in 95 TFE. For SBA monomer, atypical beta-sheet first converts to typical beta-sheet at low TFE concentration (10), which then leads to a nonnative alpha-helix at higher TFE concentration. From temperature-dependent studies (5-60 degrees C) of TFE perturbation, typical beta-sheet structure appears to be less stable than atypical beta-sheet and the induced helix entails reduced thermal stability. The heat induced transitions are reversible except for atypical to typical beta-sheet conversion. FTIR results reveal a partial alpha-helix conversion at high protein concentration but with quantitative yield. However, aggregation is detected with FTIR at lower TFE concentration, which disappears in more TFE. Near-UV CD, fluorescence and phosphorescence studies imply the existence of an intermediate with native-like secondary and tertiary structure, which could be related to the dissociation of tetramer to monomer. This has been further supported by concentration dependent far-UV CD studies. Chemical modification with N-bromosuccinimide (NBS) shows that all six tryptophans per monomer are solvent-exposed in the induced alpha-helical conformation. These results may provide novel and important insights into the perturbed folding problem of SBA in particular, and beta-sheet oligomeric proteins in general. (c) 2012 Elsevier Masson SAS. All rights reserved.