title: Mechanism of 1-Cys type methionine sulfoxide reductase A regeneration by glutaredoxin.
abstract: Glutaredoxin (Grx), a major redox regulator, can act as a reductant of methionine sulfoxide reductase A (MsrA). However, the biochemical mechanisms involved in MsrA activity regeneration by Grx remain largely unknown. In this study, we investigated the regeneration mechanism of 1-Cys type Clostridium oremlandii MsrA (cMsrA) lacking a resolving Cys residue in a Grx-dependent assay. Kinetic analysis showed that cMsrA could be reduced by both monothiol and dithiol Grxs as efficiently as by in vitro reductant dithiothreitol. Our data revealed that the catalytic Cys sulfenic acid intermediate is not glutathionylated in the presence of the substrate, and that Grx instead directly formed a complex with cMsrA. Mass spectrometry analysis identified a disulfide bond between the N-terminal catalytic Cys of the active site of Grx and the catalytic Cys of cMsrA. This mixed disulfide bond could be resolved by glutathione. Based on these findings, we propose a model for regeneration of 1-Cys type cMsrA by Grx that involves no glutathionylation on the catalytic Cys of cMsrA. This mechanism contrasts with that of the previously known 1-Cys type MsrB.
authors.affiliation: Department of Biochemistry and Molecular Biology, Yeungnam University College of Medicine, Daegu 705-717, Republic of Korea.
authors.forename: Moon-Jung
authors.initials: MJ
authors.lastname: Kim
authors.affiliation: Graduate School of Pharmaceutical Sciences, College of Pharmacy, Ewha Womans University, Seoul 120-750, Republic of Korea.
authors.forename: Jaeho
authors.initials: J
authors.lastname: Jeong
authors.affiliation: Graduate School of Pharmaceutical Sciences, College of Pharmacy, Ewha Womans University, Seoul 120-750, Republic of Korea.
authors.forename: Jihye
authors.initials: J
authors.lastname: Jeong
authors.affiliation: Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
authors.forename: Kwang Yeon
authors.initials: KY
authors.lastname: Hwang
authors.affiliation: Graduate School of Pharmaceutical Sciences, College of Pharmacy, Ewha Womans University, Seoul 120-750, Republic of Korea. Electronic address: kjl@ewha.ac.kr.
authors.forename: Kong-Joo
authors.initials: KJ
authors.lastname: Lee
authors.affiliation: Department of Biochemistry and Molecular Biology, Yeungnam University College of Medicine, Daegu 705-717, Republic of Korea. Electronic address: hykim@ynu.ac.kr.
authors.forename: Hwa-Young
authors.initials: HY
authors.lastname: Kim
mesh_terms: D000595:Amino Acid Sequence; D003013:Clostridium / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D003545:Cysteine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D054477:Glutaredoxins / Q000737:chemistry / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D007700:Kinetics; D051150:Methionine Sulfoxide Reductases / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D013434:Sulfenic Acids / Q000737:chemistry / Q000378:metabolism
chemical_list: D054477:Glutaredoxins; D013434:Sulfenic Acids; D051150:Methionine Sulfoxide Reductases; C100870:cysteinesulfenic acid; D005978:Glutathione; D003545:Cysteine
keywords: 1-Cys MsrA; Disulfide bond; Glutaredoxin; Glutathionylation; Regeneration