Analysis of the factors involved in the loss and restoration of the chaperone-like function of alpha-crystallin.

alpha-crystallin, the major protein component of the crystallin lens of mammalian eyes, is found in vivo as two separate gene products. Both isoforms are expressed in different major tissues of the body, with the lens the only location where both are found together. Both sequences can be phosphorylated, though at different locations. Both exhibit a high sequence homology to the small heat shock proteins, and it has been shown that alpha-crystallin also resists heat-induced denaturation. Horwitz [J. Horowitz (1992) Proc. Natl. Acad. Sci. USA 89, 10449-10453] demonstrated that alpha-crystallin can exhibit chaperone-like protection against heat-induced turbidity increases, and it has been suggested that this may be an in vivo function as well. However, neither isoform, when purified, shows the same overall level of chaperone-like activity as the native species, except for one phosphorylated species [M. A. M. van Boekel, S. E. A. Hoogakker, J. J. Harding, and W. W. de Jong (1996) Ophthalmic Res. 28(Suppl. 1), 32-38]. Experiments designed to determine the factors leading to loss of chaperone-like activity indicate that strong ionic conditions, such as those used in isoform separation and/or the presence of divalent cations reduce the efficiency of this function and that the presence of EDTA fully restores it irrespective of prior treatment or buffer conditions. Heat stability is essentially preserved under all conditions. These results suggest that alpha-crystallin may serve primarily as a heat shock protein in vivo and that the chaperone-like function may be inhibited under physiological conditions.