Gamma S-crystallin of bovine and human eye lens: solution structure, stability and folding of the intact two-domain protein and its separate domains

Abstract

Human and bovine γS-crystallin (HγS and BγS) and their isolated N- and C-terminal domains were cloned and expressed as recombinant proteins in E. coli. HγS and BγS are found to be authentic according to their spectral and hydrodynamic properties. Both full-length proteins and isolated domains are monomeric and exhibit high thermal and pH stabilities. The thermodynamic characterization made use of chemically and thermally-induced equilibrium unfolding transitions at varying pH. In spite of its exemplary two-domain structure, γS-crystallin does not show bimodal unfolding characteristics. In the case of BγS, at pH 7.0, the C-terminal domain is less stable than the N-terminal one, whereas for HγS the opposite holds true. Differential scanning calorimetry confirms the results of chemically-induced equilibrium unfolding transitions. Over the whole pH range between 2.0 and 11.5, HγS-crystallin and its isolated domains (HγS-N and HγS-C) follow the two-state model. The two-state unfolding of the intact two-domain protein points to the close similarity of the stabilities of the constituent domains. Obviously, interactions between the domains do not contribute significantly to the overall stability of γS-crystallin. In contrast, the structurally closely related γB-crystallin owes much of its extreme stability to domain interactions.