Heat-induced conformational change and increased chaperone activity of lens a-crystallin

Abstract

PURPOSE. Alpha-crystallin is the major structural protein of the eye lens known to have chaperone-like activity. Our objective is to elucidate the nature of the thermal transition that a-crystallin undergoes at 60°C and the effect of this transition on the chaperone activity. METHODS. FPLC size exclusion chromatography, far- and near-ultraviolet circular dichroism, and tryptophan (Trp) and 1-anilino-8-naphthalenesulfonate (ANS) fluorescence were used to study conformational change. Turbidity of dithiothreitol (DTT)-reduced insulin was used to study chaperone activity. RESULTS. The thermal transition was identified as a conformational change in mainly tertiary (partial unfolding) and quaternary high-molecular-weight (HMW) aggregation structures, along with a loss of 10 percentage points of secondary structure (ß-sheet). Initial partial perturbation in tertiary structure increased chaperone activity, but the increase was less in the HMW aggregate. Similar results were observed in in vivo -formed HMW a-crystallin. CONCLUSIONS. The conformational change and HMW aggregation of a-crystallin observed at 60°C, as well as in vivo -formed HMW aggregates, increased chaperone activity.