Studies on the alpha-crystallin target protein binding sites: sequential binding with two target proteins

PURPOSE: alpha -Crystallin belongs to a class of small heat shock proteins and is shown to prevent aggregation of several proteins. We have shown that the temperature-induced structural perturbation leads to several fold enha nced activity. The purpose of this study was to investigate the availabilit y and specificity of the hydrophobic sites that might become available at e levated temperatures. Specifically, we address the following question: Is t here an increased exposure of fixed number of hydrophobic sites as a functi on of temperature or does a new set of sites become available at elevated t emperatures? METHODS: alpha -Crystallin target protein complexes were made at two differ ent temperatures and this complex was investigated for its chaperone-like a ctivity towards the same target protein and also other target proteins. DTT -induced aggregation of insulin, alpha -lactalbumin, thermal aggregation of betaL- and gamma -crystallin, and photo-aggregation of gamma -crystallin w ere used as model systems. Increased light scattering was used to monitor t he progress of aggregation. RESULTS: alpha -Crystallin target protein complex prepared at 37 degreesC t emperature was effective against thermal aggregation of betaL-crystallin as well as non-thermal aggregation at elevated temperatures. However, the com plex prepared at high temperature was ineffective at lower temperatures as well as with other target proteins at both temperatures. CONCLUSIONS: More target protein binding sites become available at elevated temperatures. The sites available at low temperature are a subset of the t otal sites available at elevated temperatures.