PROPERTIES OF RABBIT LIVER ALDEHYDE OXIDASE AND THE RELATIONSHIP OF THE ENZYME TO XANTHINE-OXIDASE AND DEHYDROGENASE

The properties of the molybdenum iron-sulfur flavoprotein, aldehyde ox idase from rabbit livers, have been further investigated in comparison with bovine milk xanthine oxidase. In agreement with earlier work, th e ultraviolet/visible spectra indicate that the flavin and iron-sulfur centres of the enzymes are quite similar to one another. The molybden um centres have been compared by EPR spectroscopy of molybdenum(V) and regarding re-insertion of the sulfide ligand of molybdenum into the d esulfo enzyme forms. The pH optimum for sulfide insertion is approxima te to 2 lower for aldehyde oxidase than for xanthine oxidase. A detail ed comparison of molybdenum(V) EPR signals has been made for the signa ls known as Arsenite, Slow and Rapid. Computer simulation of spectra i n (H2O)-H-1 and (H2O)-H-2, at 9 and 35 GHz was used. Slow signals from the two enzymes are scarcely distinguishable from one another. Under the conditions used, aldehyde oxidase yielded only the Rapid type 2 si gnal, whereas xanthine oxidase gives both the Rapid type 1 and 2 signa ls. The nature of the structural difference between the Rapid type and type 2 signal-giving species is discussed. It is concluded that the m olybdenum centres of xanthine oxidase and aldehyde oxidase are indeed similar to one another and that such differences as exist between thei r molybdenum(V) EPR signals and re-sulfuration properties are related to differences only in the substrate-binding sites, N-terminal amino a cid analyses have been performed on peptides obtained by trypsin cleav age of aldehyde oxidase. Comparison with a sequence previously deduced [Wright, R. M., Vaitaitis, G, M., Wilson, C. M., Repine, T. B., Terad a, L. S. & Repine, J. E. (1993) Proc. Natl. Acad. Sci. USA 90, 10690-1 0694] makes it clear that the latter is not. as was assumed, that of a xanthine dehydrogenase but of an aldehyde oxidase. In contrast to the situation with xanthine oxidase, attempts to convert non-proteolysed aldehyde oxidase to a dehydrogenase form by treatment with dithiothrei tol were unsuccessful. The reason for this is considered in the light of sequence data in the literature. The location of the NAD(+)-binding site is discussed, and the sequence data are also discussed in relati on to the molybdenum, iron-sulfur and substrate-binding sites.