Heat-induced conformational change of human lens recombinant alpha A- and alpha B-crystallins

Purpose: To determine which component of lens alpha-crystallin is responsib le for heat-induced transition, conformational change and high molecular we ight (HMW) aggregation. Methods: Recombinant alpha A- and alpha B-crystallins were used. Temperatur e dependent changes were probed by Trp fluorescence and circular dichroism (CD) measurements. HMW aggregates were induced by heating at 62 degrees C f or 1-2 h and then cooling to room temperature. The nature of HMW aggregatio n was studied with fluorescent probes, 4,4'-dianilino-1,1'-binaphthalene-5, 5'-disulfonic acid (bis-ANS) and thioflavin T (ThT). Results: CD and Trp fluorescence revealed that alpha B-crystallin was more susceptible than alpha A-crystallin to heat-induced conformational change a nd aggregation. At temperatures greater than 70 degrees C, alpha B-crystall in precipitated but alpha A-crystallin remained soluble. Both bis-ANS and T hT probes displayed increased fluorescence intensity with HMW aggregation, but the increase for bis-ANS was greater with alpha B-crystallin than with alpha A-crystallin, while the reverse was true for ThT. Conclusions: These results indicate that alpha B-crystallin is more suscept ible than alpha A-crystallin to heat-induced conformational change and aggr egation and are consistent with the notion that alpha A- and alpha B-crysta llins have different biochemical and biophysical properties in spite of the ir high degree of homology.