Regeneration Mechanisms of Arabidopsis thaliana Methionine Sulfoxide Reductases B by Glutaredoxins and Thioredoxins

• Published in: The Journal of biological chemistry Vol. 284; no. 28; pp. 18963 - 18971
• Main Authors: Tarrago, Lionel, Laugier, Edith, Zaffagnini, Mirko, Marchand, Christophe, Le Maréchal, Pierre, Rouhier, Nicolas, Lemaire, Stéphane D., Rey, Pascal
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 10.07.2009; American Society for Biochemistry and Molecular Biology
• Subjects: Arabidopsis - metabolism; Biochemistry; Biochemistry, Molecular Biology; Catalysis; Cysteine - chemistry; Enzyme Catalysis and Regulation; Glutaredoxins - metabolism; Glutathione - chemistry; Kinetics; Life Sciences; Methionine Sulfoxide Reductases; Models, Biological; Mutagenesis, Site-Directed; Mutation; Oxidoreductases - chemistry; Plant Physiological Phenomena; Protein Structure, Tertiary; Regeneration; Sulfhydryl Compounds - chemistry; Thioredoxins - chemistry; PROTEIN; ACTIVE-SITE; REGENERATION; ESCHERICHIA-COLI; PEROXIREDOXIN; BIOCHEMICAL-CHARACTERIZATION; ARABIDOPSIS THALIANA; POPLAR; REDOX REGULATION; REDUCING AGENT; PEUPLIER; CATALYTIC MECHANISM; S-GLUTATHIONYLATION; METHIONINE
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M109.015487

Methionine oxidation leads to the formation of S- and R-diastereomers of methionine sulfoxide (MetSO), which are reduced back to methionine by methionine sulfoxide reductases (MSRs) A and B, respectively. MSRBs are classified in two groups depending on the conservation of one or two redox-active Cys; 2-Cys MSRBs possess a catalytic Cys-reducing MetSO and a resolving Cys, allowing regeneration by thioredoxins. The second type, 1-Cys MSRBs, possess only the catalytic Cys. The biochemical mechanisms involved in activity regeneration of 1-Cys MSRBs remain largely elusive. In the present work we used recombinant plastidial Arabidopsis thaliana MSRB1 and MSRB2 as models for 1-Cys and 2-Cys MSRBs, respectively, to delineate the Trx- and glutaredoxin-dependent reduction mechanisms. Activity assays carried out using a series of cysteine mutants and various reductants combined with measurements of free thiols under distinct oxidation conditions and mass spectrometry experiments show that the 2-Cys MSRB2 is reduced by Trx through a dithiol-disulfide exchange involving both redox-active Cys of the two partners. Regarding 1-Cys MSRB1, oxidation of the enzyme after substrate reduction leads to the formation of a stable sulfenic acid on the catalytic Cys, which is subsequently glutathionylated. The deglutathionylation of MSRB1 is achieved by both mono- and dithiol glutaredoxins and involves only their N-terminal conserved catalytic Cys. This study proposes a detailed mechanism of the regeneration of 1-Cys MSRB activity by glutaredoxins, which likely constitute physiological reductants for this type of MSR.