Proteomic analysis of the oxidation of cysteine residues in human age-related nuclear cataract lenses.

Loss of protein thiols is a key feature associated with the onset of age-related nuclear cataract (ARNC), however, little is known about the specific sites of oxidation of the crystallins. We investigated cysteine residues in ARNC lenses and compared them with age-matched normal lenses. Proteomic analysis of tryptic digests revealed ten cysteine residues in older normal lenses that showed no significant oxidation compared to foetal counterparts (Cys 170 in betaA1/3-crystallin, Cys 32 in betaA4-crystallin, Cys 79 in betaB1-crystallin, Cys 22, Cys 78/79, C153 in gammaC-crystallin and Cys 22, Cys 24 and Cys 26 in gammaS-crystallin). Although these thiols were not oxidised in normal lenses past the 6th decade, they were present largely as disulphides in the ARNC lenses. By contrast, two cysteine residues, Cys 41 in gammaC-crystallin and Cys 18 in gammaD-crystallin, were not oxidised, even in advanced ARNC lenses. These cysteines are buried deep within the protein and any unfolding associated with cataract must be insufficient to expose them to the oxidative environment present in the centre of advanced ARNC lenses. The vast majority of the loss of protein thiol observed in such lenses is due to disulphide bond formation.