Reactions of dimethylsulfoxide reductase in the presence of dimethyl sulfide and the structure of the dimethyl sulfide-modified enzyme

The bis-molybdopterin enzyme dimethylsulfoxide reductase (DMSOR) from Rhodo bacter capsulatus catalyzes the conversion of dimethyl sulfoxide (DMSO) to dimethyl sulfide (DMS), reversibly, in the presence of suitable e(-)-donors or e(-)-acceptors. The catalytically significant intermediate formed by re action of DMSOR with DMS ('the DMS species') and a damaged enzyme form deri ved by reaction of the latter with O-2 (DMS-modified enzyme, DMSORmodD) hav e been investigated. Evidence is presented that Mo in the DMS species is no t, as widely assumed, Mo(IV). Formation of the DMS species is reversed on r emoving DMS or by addition of an excess of DMSO. Equilibrium constants for the competing reactions of DMS and DMSO with the oxidized enzyme (K-d = 0.0 7 +/- 0.01 and 21 +/- 5 mM, respectively) that control these processes indi cate formation of the DMS species occurs at a redox potential that is 80 mV higher than that required, according to the literature, for reduction of M o(VI) to Mo(IV) in the free enzyme. Specificity studies show that with dime thyl selenide, DMSOR yields a species analogous to the DMS species but with the 550 nm peak blue-shifted by 27 nm. It is concluded from published redo x potential data that this band is due to metal-to-ligand charge transfer f rom Mo(V) to the chalcogenide. Since the DMS species gives no EPR signal in the normal or parallel mode, a free radical is presumed to be in close pro ximity to the metal, most likely on the S. The species is thus formulated a s Mo-V-O-S . Me-2. Existing X-ray crystallographic and Raman data are consi stent with this structure. Furthermore, le(-) oxidation of the DMS species with phenazine ethosulfate yields a Mo(V) form without an -OH ligand, since its EPR signal shows no proton splittings. This form presumably arises via dissociation of DMSO. The structure of DMSORmodD has been determined by X- ray crystallography. All four thiolate ligands and Oy of serine-147 remain coordinated to Mo, but there are no terminal oxygen ligands and Mo is Mo(VI ). Thus, it is a dead-end species, neither oxo group acceptance nor c-donat ion being possible. O-2-dependent formation of DMSORmodD represents noncata lytic breakdown of the DMS species by a pathway alternative to that in turn over, with oxidation to Mo(VI) presumably preceding product release. Steps in the forward and backward catalytic cycles are discussed in relation to e arlier stopped-flow data. The finding that in the back-assay the Mo(IV) sta te may at least in part be by-passed via two successive le- reactions of th e DMS species with the e(-)-acceptor, may have implications in relation to the existence of separate molybdopterin enzymes catalyzing DMSO reduction a nd DMS oxidation, respectively.