Exposure of bovine alpha-crystallin to 0.1 M glycine at pH 7 decreases the average molar mass of the protein from 700 to 420 kDa. When the pH is lowered to 2.5, in the same buffer, the alpha B chains specifically dissociate from the aggregates, leaving a particle of 290 kDa containing only alpha A chains. The decrease in the molar mass corresponds to the mass of the alpha B chains in the original aggregate. The pH-dependent dissociation is fully reversible. Similar changes were observed with rat and kangaroo alpha-crystallins but the dogfish protein was not affected. Sedimentation velocity analyses and fluorescence spectroscopy yielded a pK, for the dissociation, of 3.7 for alpha-crystallin and 4.0 for a homopolymer constructed from purified alpha B2 polypeptides. An alpha A2 homopolymer was virtually unaffected by the lowering of pH. The products from the dissociation were isolated and their properties studied by sedimentation analysis and acrylamide quenching of tryptophan fluorescence. The alpha B chains were found to be completely denatured, whereas the structure of the alpha A chains, in the 290 kDa, particle, were only slightly altered. Comparisons of the sequences of the various proteins examined suggested that decreased ionization of aspartic acid 127 in the alpha B chain was responsible for the specific dissociation of this polypeptide.