Immunoblot Detection of Proteins That Contain Cysteine Sulfinic or Sulfonic Acids with Antibodies Specific for the Hyperoxidized Cysteine-Containing Sequence

The cysteine residues in the active sites of certain proteins, including peroxiredoxin (Prx), Parkinson’s disease protein DJ-1, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), are more readily oxidized to sulfenic acid (Cys-SOH) than are other cysteines because of their environment. The unstable sulfenic acid undergoes further hyperoxidation to sulfinic (Cys-SO2H) and sulfonic (Cys-SO3H) acids. Hyperoxidation to sulfinic acid was recently implicated as a reversible posttranslational modification responsible for regulation of protein function. Studies of such a role have been hampered, however, by the technically demanding nature of assays for hyperoxidized proteins. Here we describe the production and characterization of antibodies to hyperoxidized peptides modeled on the active sites of Prx, DJ-1, and GAPDH. The utility of these antibodies in the study of protein function should prove similar to that of antibodies to peptides containing phosphoserine or phosphothreonine.