STRUCTURAL CHARACTERIZATION OF FOLDED AND UNFOLDED STATES OF AN SH3 DOMAIN IN EQUILIBRIUM IN AQUEOUS BUFFER

The isolated N-terminal Src homology 3 (SH3) domain of Drosophila drk exists in equilibrium between folded and unfolded states in aqueous bu ffer near neutral pH. Nuclear magnetic resonance spectra recorded on b oth states simultaneously exhibit an approximate 1:1 ratio of protein conformations. The folded form is similar to other known SH3 structure s, especially the N-terminal SH3 domain of the mammalian homologue GRB 2, A stretch of sequential amide-amide nuclear Overhauser effect cross -peaks for resonances of the unfolded state is observed in a region co rresponding to beta-strands in the folded state. The results suggest t hat turn-like conformations may be preferentially sampled in the foldi ng pathway for this predominantly beta-structured SH3 domain. In addit ion, a stable turn at Leu-28 is observed in the unfolded but not in th e folded state. Comparison of this unfolded form with a denatured stat e in 2 M guanidine hydrochloride shows that, while both are highly dis ordered, these states are not identical and more residual structure is present under nondenaturing conditions.