NIH Research Festival
Methionine sulfoxide reductase A (msrA) reduces protein methionine sulfoxides to methionine. Its catalytic cysteine (Cys72-SH) has a low pKa that facilitates oxidation of the thiol to form cysteine sulfenic acid. When the catalytic cycle proceeds efficiently, the sulfenic acid is reduced back to cysteine at the expense of thioredoxin. However, sulfenic acid is vulnerable to “irreversible” oxidation to cysteine sulfinic acid (hyperoxidation). We observed that human msrA is resistant to hyperoxidation while mouse msrA is readily hyperoxidized by micromolar concentrations of hydrogen peroxide. We found that the sulfenic acid in human msrA rapidly forms a sulfenamide with nearby Trp74 that is resistant to further oxidation. Mouse msrA is slow to form the sulfenamide, explaining its sensitivity to hyperoxidation. Slow sulfenamide formation requires the presence of Met229 in the carboxyl domain. This residue is a Val in human msrA, and mutation of Val229 to Met rendered human msrA sensitive to hyperoxidation. Human msrA remained resistant to hyperoxidation when Val229 was mutated to 5 other residues. The carboxyl domain of msrA is known to be flexible, with access to the active site. Met residues are also known to form stable, non-covalent bonds with aromatic residues through interaction of the sulfur atom with the aromatic ring. We propose that Met229 forms a bond with the aromatic ring of Trp74 at the active site, preventing formation of the sulfenamide. As a consequence, the sulfenic acid remains available for facile, irreversible oxidation to cysteine sulfinic acid.
Scientific Focus Area: Molecular Biology and Biochemistry
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