Purification and characterization of a prokaryotic xanthine dehydrogenase from Comamonas acidovorans

Biochemistry
Q Xiang, D E Edmondson

Abstract

Xanthine dehydrogenase (XDH) is induced in Comamonas acidovorans cells incubated in a limited medium with hypoxanthine as the only carbon and nitrogen source. The enzyme has been purified to homogeneity using standard techniques and characterized. It contains two subunits with M(r) values of 90 and 60 kDa. Gel filtration studies show the enzyme to have an alpha 2 beta 2 native structure. No precursor form of the enzyme is observed on Western blot analysis of cell extracts obtained at various stages of enzyme induction. Metal analysis of the purified enzyme shows 1.1 Mo, 4.0 Fe, and 3.6 phosphorus atoms per alpha beta protomer. Cofactor analysis shows the enzyme to contain a single molybdopterin mononucleotide and one FAD per alpha beta protomer. Electron spin resonance and circular dichroism spectral studies of the oxidized and reduced forms of the enzyme suggest the Fe centers to be two nonidentical [2Fe-2S] clusters. Electron spin resonance signals due to Mo(V) and neutral FAD radical are also observed in the reduced form of the enzyme. Purified enzyme preparations ranged from 70% to 100% functionality. The enzyme is irreversibly inactivated by CN- and is inhibited on incubation with allopurinol. With xanthine and NAD+ as sub...Continue Reading

References

Jun 1, 1977·Journal of Bacteriology·C A Woolfolk, J S Downard
Jul 15, 1993·European Journal of Biochemistry·A Schieber, D E Edmondson

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Citations

Apr 25, 2009·Journal of Natural Products·James M Pauff, Russ Hille
Sep 22, 1999·European Journal of Biochemistry·T SchräderJ R Andreesen
Feb 5, 2014·International Journal of Molecular Sciences·Ana Rita Otrelo-CardosoTeresa Santos-Silva
Sep 8, 2012·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Yuran ZhangWu Wang
Mar 2, 2019·Journal of Environmental Science and Health. Part A, Toxic/hazardous Substances & Environmental Engineering·Sarani Sen, Priyabrata Sarkar
Apr 9, 2008·The Journal of Biological Chemistry·Silvia SchumannSilke Leimkühler
Oct 27, 2017·Journal of Biochemical and Molecular Toxicology·Shivraj Hariram NileDoo Hwan Kim
Jul 22, 2004·The Journal of Biological Chemistry·Silke LeimkühlerRuss Hille
Nov 7, 1996·Chemical Reviews·Russ Hille

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