Membrane-Bound Flavocytochrome MsrQ Is a Substrate of the Flavin Reductase Fre in Escherichia coli

Abstract

MsrPQ is a new type of methionine sulfoxide reductase (Msr) system found in bacteria. It is specifically involved in the repair of periplasmic methionine residues that are oxidized by hypochlorous acid. MsrP is a periplasmic molybdoenzyme that carries out the Msr activity, whereas MsrQ, an integral membrane-bound hemoprotein, acts as the physiological partner of MsrP to provide electrons for catalysis. Although MsrQ (YedZ) was associated since long with a protein superfamily named FRD (ferric reductase domain), including the eukaryotic NADPH oxidases and STEAP proteins, its biochemical properties are still sparsely documented. Here, we have investigated the cofactor content of the E. coli MsrQ and its mechanism of reduction by the flavin reductase Fre. We showed by electron paramagnetic resonance (EPR) spectroscopy that MsrQ contains a single highly anisotropic low-spin (HALS) b-type heme located on the periplasmic side of the membrane. We further demonstrated that MsrQ holds a flavin mononucleotide (FMN) cofactor that occupies the site where a second heme binds in other members of the FDR superfamily on the cytosolic side of the membrane. EPR spectroscopy indicates that the FMN cofactor can accommodate a radical semiquinone species. The cytosolic flavin reductase Fre was previously shown to reduce the MsrQ heme. Here, we demonstrated that Fre uses the FMN MsrQ cofactor as a substrate to catalyze the electron transfer from cytosolic NADH to the heme. Formation of a specific complex between MsrQ and Fre could favor this unprecedented mechanism, which most likely involves transfer of the reduced FMN cofactor from the Fre active site to MsrQ.

The phylogeny of Acetobacteraceae : photosynthetic traits and deranged respiratory enzymes

“…5B ). It contains two different proteins coded by adjacent genes resembling the yedYZ of Escherichia coli , recently re-named MsrPQ for it constitutes a bacterial methionine sulfoxide reductase ( 66 , 67 ). This enzyme belongs to the group of sulfite oxidases ( 68 ) and helps in quenching oxidative damage to proteins by reducing methionine sulfoxides ( 66 , 67 ).…”

“…It contains two different proteins coded by adjacent genes resembling the yedYZ of Escherichia coli , recently re-named MsrPQ for it constitutes a bacterial methionine sulfoxide reductase ( 66 , 67 ). This enzyme belongs to the group of sulfite oxidases ( 68 ) and helps in quenching oxidative damage to proteins by reducing methionine sulfoxides ( 66 , 67 ). Although MsrPQ normally oxidizes ubiquinol to reduce methionine sulfoxides in the periplasm ( 68 ), nSor likely functions as a Q reductase because its membrane di-heme cytochrome b is structurally similar ( Fig.…”

X-Ray Structure and enzymatic study of a Bacterial NADPH oxidase highlight the activation mechanism of eukaryotic NOX

“…The strict requirement of FAD addition for SpNOX DH activity and its µM level of affinity suggests that the flavin behaves as a co-substrate rather than a prosthetic group. As an isolated domain, SpNOX DH may work as a flavin reductase enzyme (Gaudu et al, 1994; Fieschi et al, 1995; Niviere et al, 1996), making an interesting parallel with the E.coli flavin reductase fre (also in the FNR family) which reduces the MsrQ transmembrane component (in the FRD family, like the NOX TM domain, see Figure 6) (Caux et al, 2021; Juillan-Binard et al, 2017).…”

Roles of methionine and cysteine residues of the Escherichia coli sensor kinase HprS in reactive chlorine species sensing

“…6). The reduction mechanism of MsrQ by the cytoplasmic flavin reductase Fre was recently demonstrated [48], which suggests that ubiquinone is not the only electron donor for the MsrPQ system [38]. It is important to clarify the relationship among the MsrP/Q, DsbAB, and Fre systems to elucidate the molecular mechanism of HprS activation (Fig.…”