title: Changes in solvent accessibility of wild-type and deamidated βB2-crystallin following complex formation with αA-crystallin.
abstract: 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.
authors.affiliation: Oregon Health and Science University, Integrative Biosciences, 611 SW Campus Dr., Portland, OR 97239, USA. lampik@ohsu.edu
authors.forename: Kirsten J
authors.initials: KJ
authors.lastname: Lampi
authors.forename: Cade B
authors.initials: CB
authors.lastname: Fox
authors.forename: Larry L
authors.initials: LL
authors.lastname: David
mesh_terms: D002850:Chromatography, Gel; D003641:Deamination; D041961:Deuterium Exchange Measurement; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007908:Lens, Crystalline / Q000737:chemistry*; D013058:Mass Spectrometry; D010446:Peptide Fragments / Q000737:chemistry; D012997:Solvents / Q000737:chemistry*; D038202:alpha-Crystallin A Chain / Q000737:chemistry*; D038206:beta-Crystallin B Chain / Q000737:chemistry*
chemical_list: D010446:Peptide Fragments; D012997:Solvents; D038202:alpha-Crystallin A Chain; D038206:beta-Crystallin B Chain; C113634:beta-crystallin B2