Unfolding crystallins: The destabilizing role of a β‐hairpin cysteine in βB2‐crystallin by simulation and experiment

Abstract

The thermodynamic and kinetic stabilities of the eye lens family of βγ‐crystallins are important factors in the etiology of senile cataract. They control the chance of proteins unfolding, which can lead to aggregation and loss of transparency. βB2‐Crystallin orthologs are of low stability and comprise two typical βγ‐crystallin domains, although, uniquely, the N‐terminal domain has a cysteine in one of the conserved folded β‐hairpins. Using high‐temperature (500 K) molecular dynamics simulations with explicit solvent on the N‐terminal domain of rodent βB2‐crystallin, we have identified in silico local flexibility in this folded β‐hairpin. We have shown in vitro using two‐domain human βB2‐crystallin that replacement of this cysteine with a more usual aromatic residue (phenylalanine) results in a gain in conformational stability and a reduction in the rate of unfolding. We have used principal components analysis to visualize and cluster the coordinates from eight separate simulated unfolding trajectories of both the wild‐type and the C50F mutant N‐terminal domains. These data, representing fluctuations around the native well, show that although the mutant and wild‐type appear to behave similarly over the early time period, the wild type appears to explore a different region of conformational space. It is proposed that the advantage of having this low‐stability cysteine may be correlated with a subunit‐exchange mechanism that allows βB2‐crystallin to interact with a range of other β‐crystallin subunits.