FOLDING KINETICS OF CHE Y MUTANTS WITH ENHANCED NATIVE ALPHA-HELIX PROPENSITIES

摘要

in this work we study the folding kinetics of Che Y mutants in which the helical propensity of each of its five alpha-helices has been greatly enhanced by local interactions (between residues close in sequence). This constitutes an experimental test on the role of local interactions in protein folding, as well as providing new information on the details of the folding pathway of the protein Che Y. With respect to the first issue, our results show that the enhancement of helical propensities by native-like local interactions in Che Y has the following general effects: (1) the energetics of the whole Che Y folding energy landscape (folded state, intermediate, denatured state and main transition state) are affected by the enhancement of helical propensities, thus, native-like local interactions appear to have a low specificity for the native conformation; (2) our results support the idea, proposed from thermodynamic analysis of the mutants, that the denatured state under native conditions becomes more compact upon enhancement of helical propensities; (3) the rate of folding in aqueous solution decreases in all the mutants, suggesting that the optimization of the folding rate in this protein requires low secondary structure propensities. Regarding the description of the folding pathway of Che Y, we find evidence that the folding transition state of Che Y is constituted by two sub-domains with different degree of helical structure. The first includes helices 1 and 2 which are rather structured, while the second encompasses the last three helices, which are very unstructured. On the other hand, the same analysis for the folding intermediate indicates that all the five alpha-helices are, on average, rather structured. Thus, suggesting that a large structural reorganization of the last three alpha-helices must take place before folding can be completed. This conclusion indicates that the folding intermediate of Che Y is a misfolded species. (C) 1997 Academic Press Limited. [References: 38]