Evidence for the Dimerization-Mediated Catalysis of Methionine Sulfoxide Reductase A from Clostridium oremlandii

Abstract

Clostridium oremlandii MsrA (CoMsrA) is a natively selenocysteine-containing methionine-S-sulfoxide reductase and classified into a 1-Cys type MsrA. CoMsrA exists as a monomer in solution. Herein, we report evidence that CoMsrA can undergo homodimerization during catalysis. The monomeric CoMsrA dimerizes in the presence of its substrate methionine sulfoxide via an intermolecular disulfide bond between catalytic Cys16 residues. The dimeric CoMsrA is resolved by the reductant glutaredoxin, suggesting the relevance of dimerization in catalysis. The dimerization reaction occurs in a concentration- and time-dependent manner. In addition, the occurrence of homodimer formation in the native selenoprotein CoMsrA is confirmed. We also determine the crystal structure of the dimeric CoMsrA, having the dimer interface around the two catalytic Cys16 residues. A central cone-shaped hole is present in the surface model of dimeric structure, and the two Cys16 residues constitute the base of the hole. Collectively, our biochemical and structural analyses suggest a novel dimerization-mediated mechanism for CoMsrA catalysis that is additionally involved in CoMsrA regeneration by glutaredoxin.

Redox Balance Differentially Affects Biomechanics in Permeabilized Single Muscle Fibres—Active and Passive Force Assessments with the Myorobot

“…In addition, methionine sulfoxide is involved in the conversion from catalytic cysteine, in its protonated form (Cys-SH), to cysteine sulfenic acid (Cys-SOH) via a reductase (see [ 47 ], Figure 1 ) [ 47 ]. The presence of both forms then allows the formation of strong disulphide bonds that, in vitro, can only be reversed by an external reducing agent (e.g., DTT) [ 47 ].…”

“…In addition, methionine sulfoxide is involved in the conversion from catalytic cysteine, in its protonated form (Cys-SH), to cysteine sulfenic acid (Cys-SOH) via a reductase (see [ 47 ], Figure 1 ) [ 47 ]. The presence of both forms then allows the formation of strong disulphide bonds that, in vitro, can only be reversed by an external reducing agent (e.g., DTT) [ 47 ]. As such, the prolonged effect of H 2 O 2 on two of its potential targets (cysteine and methionine) would suggest a marked increase in passive restoration stress through additionally introduced disulphide bonds.…”

Biocatalytic Preparation of Chiral Sulfoxides through Asymmetric Reductive Resolution by Methionine Sulfoxide Reductase A

“…[22] Based on the reported catalytic mechanismofM srA, the reductant was neededt ob reak the intramoleculard isulfide bond of MsrA after catalysis to regenerate the enzyme. [30][31] Thus, we intendedt oe stablish a pmMsrA regeneration system (Scheme2), aimed to prepare (R)-sulfoxide efficientlyu nder high substrate concentration. As thioredoxin (Trx) was the general reductantf or MsrA regenerationi nv ivo, [31][32] we firstly tested the effect of the pmMsrA-Trx system.T he whole-cell coexpressing pmTrxa nd pmMsrA proteins were applied to reduce the rac-1a at ac ell density of 15 gL À1 .H owever,c onversion of (S)-1a was still incomplete if the substrate concentration was higher than 8mm.A fter 10 ho fr eaction, the ee of (R)-1a was only 56 %a tasubstrate concentration of 10 mm ( Figure 2).…”

Distribution of methionine sulfoxide reductases in fungi and conservation of the free-methionine-R-sulfoxide reductase in multicellular eukaryotes

“…Each had another Cys outside the conserved position of the resolving Cys in canonical fungal MsrAs. These two MsrA sequences had high similarity with the Sec-MsrAs from the bacterium Alkaliphilus oremlandii and the single-cell green alga Chlamydomonas reinhardtii, previously shown to use the Sec residue for the regeneration of their activity[400,403] (Data S2D).…”