Changes in solvent accessibility of wild-type and deamidated βB2-crystallin following complex formation with αA-crystallin

Aberrant protein interactions can lead to aggregation and insolubilization, such as occurs during cataract formation. Deamidation, a prevalent age-related modification in the lens of the eye, decreases stability of the major lens proteins, crystallins. The mechanism of deamidation altering interactions between αA-crystallin and βB2-crystallin was investigated by detecting changes in solvent accessibility upon complex formation during heating. Solvent accessibility was determined by measuring hydrogen/deuterium exchange levels of backbone amides by high-resolution mass spectrometry. Deuterium levels in wild type βB2-crystallin increased 50-60% in both domains following complex formation with αA-crystallin. This increased solvent accessibility indicated a general loosening along the backbone amides. Peptides with the greatest deuterium increases were located at the buried monomer-monomer interface, suggesting that the βB2 dimer was disrupted. The only region where the deuterium levels decreased was in βB2 peptide 123-139, containing an outside loop, and may be a potential site of interaction with αA. Mimicking deamidation at the βB2 dimer interface prevented complex formation with αA. When temperatures were lowered, an αA/βB2 Q70E/Q162E complex formed with similar solvent accessibilities as αA/WT βB2. Deamidation did not disrupt specific αA/βB2 interactions but favored aggregation before complex formation with αA. We conclude that deamidation contributes to cataract formation through destabilization of crystallins before they can be rescued by α-crystallin.

► Solvent accessibilities of βB2 and αA in complex were detected by deuterium uptake. ► Exposure of the dimer interface in βB2 suggested βB2 monomers complex with αA. ► Decreased exposure in an outside loop in βB2 suggested a site of interaction with αA. ► Accessibility in the α-crystallin domain of αA increased in complex with βB2. ► Deamidated βB2 formed a similar complex with αA as βB2, but at lower temperatures.