Structure-based analysis of the β8 interactive sequence of human αB crystallin

The functional importance of the β8 sequence (131LTITSSLS138), which is on the surface of the α crystallin core domain of human αB crystallin, was evaluated using site-directed mutagenesis. Ultraviolet circular dichroism determined that mutating the surface-exposed, nonconserved residues, Leu-131, Thr-132, Thr-134, Ser-135, Ser-136, and Ser-138 individually or in combination (αAβ8 and CEβ8), had no measurable effect on secondary and tertiary structure. Size exclusion chromatography determined the size of the complexes formed by the β8 mutants to be 6−8 subunits larger than wt αB crystallin. In chaperone assays, the protective effect of the L131S, T132A, and S135C mutants of the β8 sequence was similar to wt αB crystallin when βL crystallin and alcohol dehydrogenase were the chaperone substrates and decreased to 66% when citrate synthase was the chaperone substrate. In contrast, the chaperone activity for all three substrates was dramatically reduced for the T134K, S138A, S136H, and CEβ8 mutants. The promi...