Mammals Reduce Methionine-S-sulfoxide with MsrA and Are Unable to Reduce Methionine-R-sulfoxide, and This Function Can Be Restored with a Yeast Reductase

Abstract

Methionine is an essential amino acid in mammals at the junction of methylation, protein synthesis, and sulfur pathways. However, this amino acid is highly susceptible to oxidation, resulting in a mixture of methionine-S-sulfoxide and methionine-R-sulfoxide. Whether methionine is quantitatively regenerated from these compounds is unknown. Here we report that SK-Hep1 hepatocytes grew on methionine-S-sulfoxide and consumed this compound by import and methionine-S-sulfoxide reductase (MsrA)-dependent reduction, but methionine-R-sulfoxide reductases were not involved in this process, and methionine-R-sulfoxide could not be used by the cells. However, SKHep1 cells expressing a yeast free methionine-R-sulfoxide reductase proliferated in the presence of either sulfoxide, reduced them, and showed increased resistance to oxidative stress. Only methionine-R-sulfoxide was detected in the plasma of wild type mice, but both sulfoxides were in the plasma of MsrA knock-out mice. These results show that mammals can support methionine metabolism by reduction of methionine-S-sulfoxide, that this process is dependent on MsrA, that mammals are inherently deficient in the reduction of methionine-R-sulfoxide, and that expression of yeast free methionine-R-sulfoxide reductase can fully compensate for this deficiency.Methionine (Met) is an essential amino acid in mammals. Besides its utilization for protein synthesis, Met supports the cellular S-adenosylmethionine-dependent methylation cycle and is used for biosynthesis of cysteine and other compounds ( Fig. 1) (1-3). As a central metabolite in the global sulfur and methylation pathways, Met homeostasis is subject to exquisite regulation that tightly controls fluxes of Met-derived metabolites and Met itself (4, 5).Despite extensive information on Met regulation, one pathway critical to Met availability and homeostasis did not receive sufficient attention to date. Being a sulfur-containing amino acid, Met, along with Cys, is highly susceptible to oxidation by reactive oxygen species (ROS) 2 generated during oxidative stress and normal cellular metabolism. ROS can oxidize both free Met and Met residues in proteins, resulting in a diastereomeric mixture of Met sulfoxides: Met-S-sulfoxide (Met-SO) and Met-R-sulfoxide (Met-RO) (6, 7).To repair Met-SO and Met-RO residues in proteins, cells have evolved two families of enzymes known as Met sulfoxide reductases: MsrA that is specific for Met-SO and MsrB that specifically reduces Met-RO (8). The active sites of proteins in both Msr families are better adapted for binding Met sulfoxide residues than free Met sulfoxides (6). Although both MsrA and MsrB are capable of reducing free Met sulfoxides (9), a contribution of these enzymes to the reduction of these amino acids is not known (10).Mammals have one MsrA and three MsrB isozymes, with selenoprotein MsrB1 localized to cytosol and nucleus, MsrB2 to mitochondria, and MsrB3 to the endoplasmic reticulum (9). A single MsrA is partitioned into various cellular compartments by alternative first exon s...

Selenoproteins: Molecular Pathways and Physiological Roles

“…However, fRMsr has a very low activity with the protein-based form of methionine-R-sulfoxide, and its homologs are found only in unicellular organisms. Consistent with this observation, no fRMsr activity was detected in mammals (206).…”

Insights into Function, Catalytic Mechanism, and Fold Evolution of Selenoprotein Methionine Sulfoxide Reductase B1 through Structural Analysis*

“…3, A and B) qualitatively confirm the proposed mechanism of MsrB1 oxidation through the formation of intramolecular selenenylsulfide bridge. To monitor structural changes upon formation of selenenylsulfide bond between Cys 4 and Sec/Cys 95 , we recorded 13 C-edited NOESY and two-dimensional NOESY in 99.9% D 2 O. In the two-dimensional NOESY of the oxidized protein, two new cross-peaks compared with that of the reduced protein were observed.…”

“…In MsrBs lacking resolving Cys, the sulfenic acid intermediate can be directly reduced by thioredoxin (11,12), but the selenenic acid intermediate, at least in two tested proteins, could not (12,13). Cys 4 in selenoprotein MsrB1 is required for the thioredoxin-dependent reaction and was found to serve as a resolving Cys. The resolving Cys are absent in MsrB2 and MsrB3 (13).…”

“…Msrs are crucial repair proteins that protect cells against oxidative stress and have been implicated delaying the aging process and protecting against neurodegeneration (1,2). In addition, a new class of Msr, called fRMsr, has recently been identified (3) and characterized from Escherichia coli and Saccharomyces cerevisiae (3,4). This enzyme is specific for free Met-r-SO but is not active with protein-based Met-r-SO.…”

“…This enzyme is specific for free Met-r-SO but is not active with protein-based Met-r-SO. fRMsr also functions as an antioxidant protein (4).…”