Interactions and chaperone function of αA‐crystallin with T5P γC‐crystallin mutant

T5P γC‐crystallin mutation is associated with Coppock‐like cataract, one of the autosomal dominant congenital cataracts. It is not known why the abundant α‐crystallin cannot prevent the mutation‐related aggregation. Our previous studies indicate that the mutation changes conformation and reduces solubility and stability, but it is not known whether it is these events or the loss of interaction with other crystallins that causes the cataract. It is also not known whether the α‐crystallin can protect T5P mutant as effectively from heat‐induced aggregation as the wild‐type (WT) γC‐crystallin. To investigate the mechanism of interactions and chaperone function between αA‐ and γC‐crystallin, human αA‐crystallin and W9F mutant as well as WT γC‐crystallin and T5P mutant were cloned. Interactions between αA‐ and γC‐crystallin were studied with fluorescence resonance energy transfer (FRET), and chaperone activity was assessed by the suppression of heat‐induced aggregation of substrate proteins. Conformational changes of substrate proteins were studied by spectroscopic measurements. The results indicate that the T5P mutant showed a slightly greater FRET than WT γC‐crystallin with αA‐crystallin, and αA‐crystallin could effectively prevent both WT and T5P γC‐crystallin from heat‐induced aggregation. Spectroscopic measurements show that both αA‐crystallin and γC‐crystallin underwent only slight conformational change after chaperone binding. Together with previous results obtained with a two‐hybrid system assay of interactions between αA‐ and γC‐crystallin, the present FRET and chaperone results indicate that loss of interactions of T5P mutant with other crystallins may play a larger role than the protection afforded by chaperone‐like activity in Coppock‐like cataract.