Intrapolypeptide disulfides in human alphaA-crystallin and their effect on chaperone-like function.

We report studies on the role of protein-protein disulfides (PSSP) in the age-related loss of chaperone activity of alpha-crystallins. AlphaL-crystallin fraction was isolated from human lenses of different ages and the chaperone-like activity was determined before and after treatment with glutathione reductase (GR) and NADPH. The results confirmed an age-dependent decrease in chaperone-like function and significant improvement of this function by GR treatment. Electrospray ionization mass spectrometric (ESIMS) analysis of alphaA-crystallin suggested the presence of very little protein-glutathione mixed disulfides. ESIMS analysis of Asp-N digests of alphaA-crystallin revealed that nearly all the remaining portion of Cys-131 and Cys-142 of alphaA-crystallin was present in the form of intrapolypeptide disulfide bonds. These results show for the first time that predominantly disulfide bonds formed during aging contribute to the age-dependent loss in chaperone activity of alpha-crystallin in human lenses.