In the course of aging and cataract formation, loss of lens transparency is associated with aggregation of structural lens proteins (crystallins) and increased light scattering. Many post- translational changes have been observed in both human and bovine proteins of aging and cataractous lenses. They include phosphorylation, oxidation of tryptophan, oxidation of methionine, selective degradation of polypeptide chains, disulfide bonding (protein-protein, or mixed disulfide) and non-enzymatic glycosylation. However, it is not known whether these modifications induce protein conformational change and precede protein aggregation and insolubilization. In this proposal, physicochemical measurements (UV-VIS absorption, circular dichroism, fluorescence and laser light scattering) will be employed, to observe (1) whether protein conformations (including secondary and tertiary structures, microenvironments of aromatic amino acids and sulfhydryl groups) have changed as a result of post-translational modifications, and if they do (2) whether conformationally altered lens proteins from aging and cataractous lenses give greater scattering and (3) whether these proteins become more vulnerable to exogeneous stresses (such as UV irradiation and hydrogen peroxide), (4) whether the change in protein conformation has facilitated protein aggregation and insolubilization, and finally (5) whether enzyme phosphofructokinase also undergoes similar conformational and post-translational changes as occurring in crystallins. The significance of these post-translational changes in lens protein conformation is that its may increase light scattering either through a decreased particle mobility, or through increased protein aggregation and isolubilization. The long-term objective of this proposal is to correlate post-translationally conformational changes to changes in light scattering observed both in protein solutions and in intact lenses in vitro.
| Ellis, D Z; Nathanson, J A; Rabe, J et al. (2001) Carbachol and nitric oxide inhibition of Na,K-ATPase activity in bovine ciliary processes. Invest Ophthalmol Vis Sci 42:2625-31 |
