Deamidation of the human eye lens protein γS-crystallin accelerates oxidative aging

Abstract

Cataract disease, a clouding of the eye lens due to precipitation of lens proteins, affects millions of people every year worldwide. The proteins that comprise the lens, the crystallins, show extensive post-translational modifications (PTMs) in aged and cataractous lenses, most commonly deamidation and oxidation. Although surface-exposed glutamines and asparagines show the highest rates of deamidation, multiple modifications can accumulate over time in these long-lived proteins, even for buried residues. Both deamidation and oxidation have been shown to promote crystallin aggregation in vitro; however, it is not clear precisely how these modified crystallins contribute to insolubilization. Here, we report six novel crystal structures of a major human lens protein, {gamma}S-crystallin ({gamma}S): one of the wild-type in a monomeric state, and five of deamidated {gamma}S variants, ranging from three to nine deamidation sites, after varying degrees of sample aging. Consistent with previous work that focused on single-to triple-site deamidation, the deamidation mutations do not appear to drastically change the fold of {gamma}S; however, increasing deamidation leads to accelerated oxidation and disulfide bond formation. Successive addition of deamidated sites progressively destabilized protein structure as evaluated by differential scanning fluorimetry. Light scattering showed the deamidated variants display an increased propensity for aggregation compared to the wild-type protein. The results suggest the deamidated variants are useful as models for accelerated aging; the structural changes observed over time provide support for redox activity of {gamma}S-crystallin in the human lens. HighlightsO_LINovel structures of cataract-associated variants of human eye lens protein {gamma}S-crystallin reported C_LIO_LIIncreasing deamidation of {gamma}S-crystallin decreases stability and affects aggregation propensity C_LIO_LIOverall fold of {gamma}S-crystallin maintained among deamidated and disulfide-bonded variants C_LIO_LIDeamidated {gamma}S variants form disulfide bonds more rapidly than wild-type {gamma}S C_LIO_LIPotential functional advantage of disulfide bonding in the CXCXC motif proposed C_LI