Functional Analysis of Free Methionine-R-sulfoxide Reductase from Saccharomyces cerevisiae

Abstract

Methionine sulfoxide reductases (Msrs) are oxidoreductases that catalyze thiol-dependent reduction of oxidized methionines. MsrA and MsrB are the best known Msrs that repair methionine-S-sulfoxide (Met-S-SO) and methionine-R-sulfoxide (Met-R-SO) residues in proteins, respectively. In addition, an Escherichia coli enzyme specific for free Met-R-SO, designated fRMsr, was recently discovered. In this work, we carried out comparative genomic and experimental analyses to examine occurrence, evolution, and function of fRMsr. This protein is present in single copies and two mutually exclusive subtypes in about half of prokaryotes and unicellular eukaryotes but is missing in higher plants and animals. A Saccharomyces cerevisiae fRMsr homolog was found to reduce free Met-R-SO but not free Met-S-SO or dabsyl-Met-R-SO. fRMsr was responsible for growth of yeast cells on Met-R-SO, and the double fRMsr/MsrA mutant could not grow on a mixture of methionine sulfoxides. However, in the presence of methionine, even the triple fRMsr/ MsrA/MsrB mutant was viable. In addition, fRMsr deletion strain showed an increased sensitivity to oxidative stress and a decreased life span, whereas overexpression of fRMsr conferred higher resistance to oxidants. Molecular modeling and cysteine residue targeting by thioredoxin pointed to Cys 101 as catalytic and Cys 125 as resolving residues in yeast fRMsr. These residues as well as a third Cys, resolving Cys 91 , clustered in the structure, and each was required for the catalytic activity of the enzyme. The data show that fRMsr is the main enzyme responsible for the reduction of free Met-R-SO in S. cerevisiae.

Selenoproteins: Molecular Pathways and Physiological Roles

“…Recently, in some organisms, an additional Msr was discovered, designated fRMsr, which can reduce free methionine-R-sulfoxide (204,221). 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.…”

The Response to Heat Shock and Oxidative Stress inSaccharomyces cerevisiae

“…fRMsr is thought to be the main enzyme responsible for the reduction of free Met-R-SO, whereas MsrA and MsrB are active with Met-S-SO and Met-R-SO in proteins. A triple fRMsr/ MsrA/MsrB mutant is viable on media containing methionine, but cannot grow if methionine is substituted with Met-SO (Le et al 2009). …”

“…This is particularly important because it means that methionine oxidation is readily reversible and can play an antioxidant role in scavenging ROS . Yeast contains three MSR enzymes that are required for resistance against oxidative stress (fRMsr/MsrA/MsrB) (Le et al 2009). fRMsr is thought to be the main enzyme responsible for the reduction of free Met-R-SO, whereas MsrA and MsrB are active with Met-S-SO and Met-R-SO in proteins.…”

Proteome and fermentative parameters of Saccharomyces cerevisiae CAT-1 under Very High Gravity Fermentation (VHGF) using sugarcane juice

“…This enzyme is related to both the peptidyl-prolyl isomerization during protein folding or conformational changes and the meiosis to promote an efficient sporulation when associated with other two proteins from the histone deacetylase complexes and in the glucose-stimulated transport of fructose-1,6-bisphosphatase into Vid vesicles [38,39]. The protein Mxr1-Trx2 is a methionine S-sulfoxide reductase that uses thioredoxin as electron donor [40,41]. Both thioredoxin-1 (Trx1) and thioredoxin-2 (Trx2) were previously identified in the quantitative proteome analysis of S. cerevisiae CAT-1, constituting important elements in the stress tolerance observed for this yeast [2].…”