Evidence for an essential arginine in the flavoprotein nitroalkane oxidase

G. Gadda, A. Banerjee, G. S. Fleming, P. F. Fitzpatrick

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

The flavoprotein nitroalkane oxidase from the fungus Fusarium oxysporum catalyzes the oxidative denitrification of primary or secondary nitroalkanes to yield the respective aldehydes or ketones, hydrogen peroxide and nitrite. The enzyme is inactivated in a time-dependent fashion upon treatment with the arginine-directed reagents phenylglyoxal, 2,3-butanedione, and cyclohexanedione. The inactivation shows first order kinetics with all reagents. Valerate, a competitive inhibitor of the enzyme, fully protects the enzyme from inactivation, indicating that modification is active site directed. The most rapid inactivation is seen with phenylglyoxal, with a kinact of 14.3 ± 1.1 M-1 min-1 in phosphate buffer at pH 7.3 and 30°C. The lack of increase in the enzymatic activity of the phenylglyoxal-inactivated enzyme after removing the unreacted reagent by gel filtration is consistent with inactivation being due to covalent modification of the enzyme. A possible role for an active site arginine in substrate binding is discussed.

Original languageEnglish (US)
Pages (from-to)157-163
Number of pages7
JournalJournal of Enzyme Inhibition
Volume16
Issue number2
DOIs
StatePublished - 2001

Keywords

  • Active site
  • Arginine
  • Chemical modification
  • Flavoprotein
  • Flavoprotein nitroalkane oxidase
  • Nitroalkanes
  • Phenylglyoxal

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine

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