Xanthine dehydrogenase AtXDH1 from Arabidopsis thaliana is a potent producer of superoxide anions via its NADH oxidase activity.
Xanthine dehydrogenase AtXDH1 from Arabidopsis thaliana is a key enzyme in purine degradation where it oxidizes hypoxanthine to xanthine and xanthine to uric acid. Electrons released from these substrates are either transferred to NAD(+) or to molecular oxygen, thereby yielding NADH or superoxide, respectively. By an alternative activity, AtXDH1 is capable of oxidizing NADH with concomitant formation of NAD(+) and superoxide. Here we demonstrate that in comparison to the specific activity with xanthine as substrate, the specific activity of recombinant AtXDH1 with NADH as substrate is about 15-times higher accompanied by a doubling in superoxide production. The observation that NAD(+) inhibits NADH oxidase activity of AtXDH1 while NADH suppresses NAD(+)-dependent xanthine oxidation indicates that both NAD(+) and NADH compete for the same binding-site and that both sub-activities are not expressed at the same time. Rather, each sub-activity is determined by specific conditions such as the availability of substrates and co-substrates, which allows regulation of superoxide production by AtXDH1. Since AtXDH1 exhibits the most pronounced NADH oxidase activity among all xanthine dehydrogenase proteins studied thus far, our results imply that in particular by its NADH oxidase activity AtXDH1 is an efficient producer of superoxide also in vivo.
Zarepour
Maryam
M
Membrane Protein Research Group, Department of Biochemistry, University of Alberta, 474 Medical Sciences Building, Edmonton, Alberta, T6G 2H7, Canada.
Kaspari
Katrin
K
Stagge
Stefan
S
Rethmeier
Ralf
R
Mendel
Ralf R
RR
Bittner
Florian
F
D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D009243:NAD / Q000378:metabolism*; D010084:Oxidation-Reduction; D010843:Pichia / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*; D019820:Xanthine / Q000378:metabolism; D014968:Xanthine Dehydrogenase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D011994:Recombinant Proteins; D009243:NAD; D013481:Superoxides; D019820:Xanthine; D014968:Xanthine Dehydrogenase