The reactivity of chicken liver xanthine dehydrogenase with molecular oxygen

T. Nishino, T. Nishino, Lawrence M Schopfer, V. Massey

Research output: Contribution to journalArticle

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Abstract

The reaction of 6-electron reduced chicken liver xanthine dehydrogenase (XDH) with molecular oxygen was studied using both stopped flow and steady-state turnover techniques at pH 7.8, 4°C. Oxidation of fully reduced XDH proceeded via four phases, three of which were detected with the stopped flow spectrophotometer. The fastest phase was second order in oxygen (1900 M-1 s-1), resulted in the appearance of flavin semiquinone and yielded no superoxide. The next phase was also second order in oxygen (260 M-1 s-1), involved the loss of flavin semiquinone and yielded, on average, 1 mol of superoxide/mol of XDH oxidized. The last 2 electron equivalents were located in the iron-sulfur centers. They were released one equivalent at a time in the form of superoxide. Steady-state kinetics were found to be critically dependent on temperature and oxygen concentration. When these factors were carefully controlled, both the xanthine-oxygen and NADH-oxygen reductase reactions gave linear Linewaver-Burk plots. The xanthine-oxygen data yielded a turnover number of 43 min-1, which was 42% of that for xanthine-NAD turnover. During turnover, with xanthine and O2, 40-44% of the electron equivalents introduced by xanthine appeared as superoxide. Reduced pyridine nucleotides, NAD and 3-aminopyridine adenine dinucleotide, dramatically reduced the formation of superoxide at levels which did not seriously inhibit oxygen reactivity.

Original languageEnglish (US)
Pages (from-to)2518-2527
Number of pages10
JournalJournal of Biological Chemistry
Volume264
Issue number5
StatePublished - Jan 1 1989

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Xanthine Dehydrogenase
Molecular oxygen
Liver
Xanthine
Chickens
Oxygen
Superoxides
NAD
Electrons
Spectrophotometers
Sulfur
Oxidoreductases
Nucleotides
Iron
Oxidation
Kinetics
Temperature

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

The reactivity of chicken liver xanthine dehydrogenase with molecular oxygen. / Nishino, T.; Nishino, T.; Schopfer, Lawrence M; Massey, V.

In: Journal of Biological Chemistry, Vol. 264, No. 5, 01.01.1989, p. 2518-2527.

Research output: Contribution to journalArticle

Nishino, T. ; Nishino, T. ; Schopfer, Lawrence M ; Massey, V. / The reactivity of chicken liver xanthine dehydrogenase with molecular oxygen. In: Journal of Biological Chemistry. 1989 ; Vol. 264, No. 5. pp. 2518-2527.
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