Aggregation of deamidated human βB2-crystallin and incomplete rescue by α-crystallin chaperone

Abstract

Aging of the lens is accompanied by extensive deamidation of the lens specific proteins, the crystallins. Deamidated crystallins are increased in the insoluble proteins and may contribute to cataracts. Deamidation has been shown in vitro to alter the structure and decrease the stability of human lens βB1, βB2 and βA3-crystallin. Of particular interest, βB2 mutants were constructed to mimic the effect of in vivo deamidations at the interacting interface between domains, at Q70 in the N terminal domain and at Q162, its C-terminal homologue. The double mutant was also constructed. We previously reported that deamidation at the critical interface sites decreased stability, while preserving the dimeric 3D structure. In the present study, dynamic light scattering, differential scanning calorimetry and small angle X-ray scattering were used to investigate the effect of deamidation on stability, thermal unfolding and aggregation. The bovine βLb fraction was used for comparative analysis. The chaperone requirements of the various samples were determined using bovine α-crystallins as the chaperone. Deamidation at both interface Gln residues or at Q70, but not Q162, significantly lowered the temperature for unfolding and aggregation, which was rapidly followed by precipitation. This deamidation-induced aggregation and precipitation was not completely prevented by α-crystallin chaperone. A potential mechanism for cataract formation in vivo involving accumulation of deamidated β-crystallin aggregates is discussed.