MULTIPLE STATES OF THE MOLYBDENUM CENTER OF DIMETHYLSULFOXIDE REDUCTASE FROM RHODOBACTER-CAPSULATUS REVEALED BY ER SPECTROSCOPY

The dimethylsulphoxide reductase of Rhodobacter capsulatus contains a pterin molybdenum cofactor molecule as its only prosthetic group. Kine tic studies were consistent with re-oxidation of the enzyme being rate limiting in the turnover of dimethylsulphoxide in the presence of the benzyl viologen radical. EPR spectra of molybdenum(V) were generated by reducing the highly purified enzyme under a variety of conditions, and with careful control it was possible to generate at least five cle arly distinct EPR signals. These could be simulated, indicating that e ach corresponds to a single chemical species. Structures of the signal -giving species are discussed in light of the EPR parameters and of in formation from the literature. Three of the signals show coupling of m olybdenum to an exchangeable proton and, in the corresponding species, the metal is presumed to bear a hydroxyl ligand. One signal with g(av ) 1.96 shows a very strong similarity to a signal for the desulpho for m of xanthine oxidase, while two others with g(av) values of 1.98 show a distinct similarity to signals from nitrate reductase of Escherichi a coli. These data indicate an unusual flexibility in the active site of dimethylsulphoxide reductase, as well as emphasising structural sim ilarities between molybdenum enzymes bearing different forms of the pt erin cofactor. Interchange among the different species must involve ei ther a change of coordination geometry, a ligand exchange, or both. Th e latter may involve replacement of an amino acid residue co-ordinatin g molybdenum via O or N, for a cysteine co-ordinating via S. Since the two signals with g(av) 1.96 were obtained only under specific conditi ons of reduction of the enzyme by dithionite, it is postulated that th eir generation may be triggered by reduction of the pteridine of the m olybdenum cofactor from a dihydro state to the tetrahydro state.