期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2004
卷号：101
期号：21
页码：7954-7959
DOI：10.1073/pnas.0400550101
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Through a mutagenic investigation of Gly-48, a highly conserved position in the Src homology 3 domain, we have discovered a series of amino acid substitutions that are highly destabilizing, yet dramatically accelerate protein folding, some up to 10-fold compared with the wild-type rate. The unique folding properties of these mutants allowed for accurate measurement of their folding and unfolding rates in water with no denaturant by using an NMR spin relaxation dispersion technique. A strong correlation was found between {beta}-sheet propensity and the folding rates of the Gly-48 mutants, even though Gly-48 lies in an unusual non-{beta}-strand backbone conformation in the native state. This finding indicates that the accelerated folding rates of the Gly-48 mutants are the result of stabilization of a nonnative {beta}-strand conformation in the transition-state structure at this position, thus providing the first, to our knowledge, experimentally elucidated example of a mechanism by which folding can occur fastest through a nonnative conformation. We also demonstrate that residues that are most stabilizing in the transition-state structure are most destabilizing in the native state, and also cause the greatest reductions in in vitro functional activity. These data indicate that the unusual native conformation of the Gly-48 position is important for function, and that evolutionary selection for function can result in a domain that folds at a rate far below the maximum possible.
