Purification and Properties of Xanthine Dehydrogenase from Clostridium cylindrosporum

Clostridium cylindrosporumferments xanthine, uric acid, 6,8dihydroxypurine, and other purines (6,7, 42).The initial attack on the purine structure occurs by hydrolysis of the bond between atoms 1 and 6 of xanthine, forming 4-ureido-5-imidazole carboxylic acid (43).Xanthine is the only purine known to undergo this hydrolytic cleavage.To be degraded the other purines are apparently converted to xanthine (10, 42, 44, 45).Thus, uric acid is degraded to nonpurine products by unfractionated cell extracts only under conditions favorable for its reduction (42, 44).Because of its rapid degradation, the actual reduction product has not been identified.Uric acid reduction and xanthine osidation by unfractionated extracts of Cl. cylindrosporum have both been attributedto the activity of a xanthine dehydrogenase (7,42).This enzyme may also form xanthine from other purines such as hypoxanthine and 6,8-dihydroxypurine which are degraded by Cl. cylindrosporum (7, 42).In some respects the substrate specificity of this enzyme appears to differ from that of milk xanthine osidase (11,21).However, at least a partial purification of the xanthineoxidizing enzyme from Cl. cylindrosporum is required to obtain convincing evidence on the identity of its substrates and products.Therefore, a more thorough study of the clostridial xanthine-oxidizing enzyme was undertaken.In this paper a method is described for the purification of the enzyme, and data are given on its substrate specificity, on some of its chemical and kinetic properties, and on the positions oxidized or reduced in purine conversions.