No abstract

Cataracts reduce vision in 50% of individuals over 70 years of age and are a common form of blindness worldwide. Cataracts are caused when damage to the major lens crystallin proteins causes their misfolding and aggregation into insoluble amyloids. Using a thermal stability assay, we identified a class of molecules that bind α-crystallins (cryAA and cryAB) and reversed their aggregation in vitro. The most promising compound improved lens transparency in the R49C cryAA and R120G cryAB mouse models of hereditary cataract. It also partially restored solubility in aged mouse and human lenses. These findings suggest an approach to treating cataracts by stabilizing α-crystallins.

One of the major protein components of the ocular lens, ␣-crystallin, is composed of ␣A and ␣B chain subunits that have structural homology to the family of mammalian small heat shock proteins. Like other small heat shock proteins, ␣-crystallin subunits associate to form large oligomeric aggregates that express chaperone-like activity, as defined by the ability to suppress nonspecific aggregation of proteins destabilized by treatment with a variety of denaturants including heat, UV irradiation, and chemical modification. It has been proposed that age-related loss of sequences at the C terminus of the ␣A chain subunit may be a factor in the pathogenesis of cataract due to diminished capacity of the truncated crystallin to protect against nonspecific aggregation of lens proteins. To evaluate the functional consequences of ␣-crystallin modification, two mutant forms of ␣A subunits were prepared by site-directed mutagenesis. Like wild type (WT), aggregates of ϳ540 kDa were formed from a tryptophan-free ␣A mutant (W9F). When added in stoichiometric amounts, both WT and W9F subunits completely suppressed the heat-induced aggregation of aldose reductase. In contrast, subunits encoded by a truncation mutant in which the Cterminal 17 residues were deleted (R157STOP), despite having spectroscopic properties similar to WT, formed much larger aggregates with a marked reduction in chaperone-like activity. Similar results were observed when the chaperone-like activity was assessed through inhibition of ␥-crystallin aggregation induced by singlet oxygen. These results demonstrate that the structurally conservative substitution of Phe for Trp-9 has a negligible effect on the functional interaction of ␣A subunits, and that deletion of C-terminal sequences from the ␣A subunit results in substantial loss of chaperone-like activity, despite overall preservation of secondary structure.The major components of the mammalian lens fiber cells are the ␣-, ␤-, and ␥-crystallins, which constitute an estimated 35% wet weight of the lens. The crystallins contribute to the transparency and refractive power of the lens by short range interactions among themselves and cytoskeletal elements in a highly concentrated matrix (1-3). ␣-Crystallin is one of the most abundant of the crystallins in mature lens fiber cells. It is a M r ϳ0.6 -1.0 ϫ 10 6 complex composed of two structurally related subunits, designated ␣A and ␣B, which are encoded by genes localized to chromosomes 21 and 11, respectively (4, 5).