γD-crystallin is one of the major structural proteins in human eye lens. The solubility and stability of γD-crystallin play a crucial role in maintaining the optical properties of the lens during the life span of an individual. Previous study has shown that the inherited mutation G61C results in autosomal dominant congenital cataract. In this research, we studied the effects of the G61C mutation on γD-crystallin structure, stability and aggregation via biophysical methods. CD, intrinsic and extrinsic fluorescence spectroscopy indicated that the G61C mutation did not affect the native structure of γD-crystallin. The stability of γD-crystallin against heat- or GdnHCl-induced denaturation was significantly decreased by the mutation, while no influence was observed on the acid-induced unfolding. The mutation mainly affected the transition from the native state to the intermediate but not that from the intermediate to the unfolded or aggregated states. At high temperatures, both proteins were able to form aggregates, and the aggregation of the mutant was much more serious than the wild type protein at the same temperature. At body temperature and acidic conditions, the mutant was more prone to form amyloid-like fibrils. The aggregation-prone property of the mutant was not altered by the addition of reductive reagent. These results suggested that the decrease in protein stability followed by aggregation-prone property might be the major cause in the hereditary cataract induced by the G61C mutation.

β/γ-Crystallins, the major structural proteins in human lens, are highly conserved in their tertiary structures but distinct in the quaternary structures. The N- and C-terminal extensions have been proposed to play a crucial role in mediating the size of β-crystallin assembly. In this research, we investigated the molecular mechanism underlying the congenital hereditary cataract caused by the recently characterized A2V mutation in βB2-crystallin. Spectroscopic experiments indicated that the mutation did not affect the secondary and tertiary structures of βB2-crystallin. The mutation did not affect the formation of βB2/βA3-crystallin heteromer as well as the stability and folding of the heteromer, suggesting that the mutation might not interfere with the protein interacting network in the lens. However, the tetramerization of βB2-crystallin at high protein concentrations was retarded by the A2V mutation. The mutation slightly decreased the thermal stability and promoted the thermal aggregation of βB2-crystallin. Although it did not influence the stability of βB2-crystallin against denaturation induced by chemical denaturants and UV irradiation, the A2V mutant was more prone to be trapped in the off-pathway aggregation process during kinetic refolding. Our results suggested that the A2V mutation might lead to injury of lens optical properties by decreasing βB2-crystallin stability against heat treatment and by impairing βB2-crystallin assembly into high-order homo-oligomers.