A sulfenic acid enzyme intermediate is involved in the catalytic mechanismof peptide methionine sulfoxide reductase from Escherichia coli

Methionine oxidation into methionine sulfoxide is known to be involved in m any pathologies and to exert regulatory effects on proteins. This oxidation can be reversed by a ubiquitous monomeric enzyme, the peptide methionine s ulfoxide reductase (MsrA), whose activity in vivo requires the thioredoxin- regenerating system. The proposed chemical mechanism of Escherichia coil Ms rA involves three Cys residues (positions 51, 198, and 206), A fourth Cys ( position 86) is not important for catalysis, In the absence of a reducing s ystem, 2 mol of methionine are formed per mole of enzyme for wild type and Cys->86 Ser mutant MsrA, whereas only 1 mol is formed for mutants in which either Cys-198 or Cys-206 is mutated, Reduction of methionine sulfoxide is shown to proceed through the formation of a sulfenic acid intermediate, Thi s intermediate has been characterized by chemical probes and mass spectrome try analyses. Together, the results support a three-step chemical mechanism in vivo: 1) Cys-51 attacks the sulfur atom of the sulfoxide substrate lead ing, via a rearrangement, to the formation of a sulfenic acid intermediate on Cys-BI and release of 1 mol of methionine/mol of enzyme; 2) the sulfenic acid is then reduced via a double displacement mechanism involving formati on of a disulfide bond between Cys-51 and Cys-198, followed by formation of a disulfide bond between Cys-198 and Cys-206, which liberates Cys-51, and 3) the disulfide bond between Cys-198 and Cys-206 is reduced by thioredoxin -dependent recycling system process.