THE MOLECULAR CHAPERONE ALPHA-CRYSTALLIN INHIBITS UV-INDUCED PROTEIN AGGREGATION

Solutions of gamma-crystallin, and various enzymes, at neutral pH and 24-26 degrees C, became turbid upon exposure to UV radiation at 295 or 308 nm, SDS-PAGE analysis revealed interchain cross-linking and aggre gate formation compared to dark control solutions as reported previous ly. When alpha-crystallin was added to the protein solutions in stoich iometric amounts, UV irradiation resulted in significantly less turbid ity than in the absence of alpha-crystallin. For example, addition of 0.5 mg of alpha-crystallin to 0.5 mg of gamma-crystallin in 1.0 ml sol ution yielded only 25% of the turbidity seen in the absence of alpha-c rystallin. Addition of 2.0 mg of alpha-crystallin resulted in 20% of t he turbidity. Given the molecular weights of alpha- and gamma-crystall in (about 800 kDa and 20 kDa, respectively), a gamma/alpha 1:1 weight ratio corresponds to a 40:1 molar ratio, and a gamma/alpha 1:4 weight ratio corresponds to a 10:1 molar ratio. Hence, the molar ratio of alp ha-crystallin needed to effectively protect gamma-crystallin from phot ochemical opacification was gamma/alpha = n:1, where n was in the rang e 10-40. In terms of subunits, this ratio is gamma/alpha = 1:m, where m = 1-4. Thus, each gamma-crystallin molecule needs 1-4 alpha subunits for protection. Similar stoichiometries were observed for protection of the other proteins studied. The protection stems in part from scree ning of UV radiation by alpha-crystallin but more importantly from a c haperone effect analogous to that seen in thermal aggregation experime nts.