GEOMETRY OF THE SOLUBLE METHANE MONOOXYGENASE CATALYTIC DIIRON CENTERIN 2 OXIDATION-STATES

Background: The hydroxylase component of soluble methane monooxygenase (sMMO) contains a dinuclear iron center responsible for the oxidation of methane to methanol. As isolated, the center is in the oxidized, d iiron(III) state. The 2.2 Angstrom resolution X-ray structure of the o xidized hydroxylase, H-OX, from Methylococcus capsulatus (Bath) was pr eviously determined at 4 degrees C. In this structure the two iron ato ms are bridged by a glutamate, a hydroxide ion, and an acetate ion, an d additionally coordinated to two His residues, three Glu residues, an d a water molecule. Results: The 1.7 Angstrom resolution crystal struc tures of the sMMO hydroxylase from Methylococcus capsulatus (Bath) in both its oxidized diiron(III), H-OX, and dithionite-treated, reduced d iiron(II), H-red, oxidation states were determined at -160 degrees C. The structure of the diiron center in H-OX differs from that previousl y reported at 2.2 Angstrom resolution and 4 degrees C. Although the hy droxide bridge is retained, the bidentate, bridging ligand assigned as acetate is replaced by a weakly coordinating monoatomic water bridge. In the resulting four-membered Fe(OH)Fe(OH2) ring, the Fe ... Fe dist ance is shortened from 3.4 Angstrom to 3.1 Angstrom. In protomer A of H-red, the hydroxide bridge is displaced by an oxygen atom of Glu243, which undergoes a carboxylate shift from its terminal monodentate bind ing mode in H-OX to a mode in which the carboxylate is both monoatomic bridging and bidentate chelating. We therefore conclude that the cent er has been reduced to the diiron(II) oxidation state. Both iron atoms are coordinated to five ligands and weakly to a sixth water molecule in the resulting structure. The diiron center in protomer B of H-red h as the same composition as those in H-OX. In both the oxidized and red uced structures, the diiron core is connected through hydrogen bonds i nvolving exogenous species to Thr213 in the active site cavity. Conclu sions: The diiron center in H-OX can change its exogenous ligand coord ination and geometry, a property that could be important in the cataly tic cycle of sMMO. In H-red, a carboxylate shift occurs, extruding hyd roxide ion and opening coordination sites for reaction with O-2 to for m the diiron(III) peroxo intermediate, H-peroxo. Residue Thr213 may fu nction in catalysis.