alpha-Crystallin, a multimeric protein present in the eye lens, is shown to have chaperone-like activity in preventing thermally induced aggregation of enzymes and other crystallins. We have studied the rapid refolding of alpha-crystallin, and compared it with other calf eye lens proteins, namely beta- and gamma-crystallins. alpha-Crystallin forms a clear solution upon rapid refolding from 8 M urea. The refolded alpha-crystallin has native-like secondary, tertiary, and quaternary structures as revealed by circular dichroism and fluorescence characteristics as well as gel filtration and sedimentation velocity measurements. On rapid refolding, beta- and gamma-crystallins aggregate and form turbid solutions. The presence of alpha-crystallin in the refolding buffer marginally increases the recovery of beta- and gamma-crystallins in the soluble form. However, unfolding of these crystallins together with alpha-crystallin using 8 M urea and subsequent refolding significantly increases the recovery of these proteins in the soluble form. These results indicate that an intermediate of alpha-crystallin formed during refolding is more effective in preventing the aggregation of beta- and gamma-crystallins. This supports our earlier hypothesis (Raman, B., and Rao, C. M. (1994) J. Biol. Chem. 269, 27264-27268) that the chaperone-like activity of alpha-crystallin is more pronounced in its structurally perturbed state.