X-ray absorption spectroscopy of xanthine oxidase. The molybdenum centres of the functional and the desulpho forms.

X-ray absorption spectra have been recorded for the molybdenum K-edge region of xanthine oxidase. Both the absorption edge and the extended fine structure (e.x.a.f.s.) regions were investigated. Spectra were obtained for samples of the desulpho enzyme as well as for mixtures of this with the active enzyme. The spectrum of the pure active form was then obtained by difference. The desulpho enzyme shows a pronounced step in the absorption edge, of a type previously associated terminal oxygen ligands. In the active enzyme this step has decreased markedly. Satisfactory simulations of the e.x.a.f.s. spectrum of the desulpho enzyme could be obtained by assuming the molybdenum to be bonded to two terminal oxygen atoms (Mo = O about .175 nm), two sulphur atoms (presumably from cysteine residues, Mo-S about .0250 nm) and one sulphur atom (presumably from a methionine residue, Mo-S about 0.290 nm). E.x.a.f.s. of the active enzyme differed appreciably from this. In keeping with earlier proposals [Gutteridge, Tanner & Bray (1978) Biochem. J. 175, 887-897], the spectrum of the active enzyme could be simulated if a sulphur atom at about 0.225 nm (i.e. presumably a terminal sulphur atom) replaced one of the terminal oxygen atoms of the desulpho from, with small changes in the other bond distances. Validity of the interpretative procedures, which involved phase shift and amplitude calculations ab initio, was demonstrated by using low molecular weight compounds of known structure.