F420H2 Oxidase (FprA) From Methanobrevibacter Arboriphilus, a Coenzyme F420-dependent Enzyme Involved in O2 Detoxification

Arch Microbiol. 2004 Oct;182(2-3):126-37. doi: 10.1007/s00203-004-0675-3. Epub 2004 Aug 31.

Abstract

Cell suspensions of Methanobrevibacter arboriphilus catalyzed the reduction of O(2) with H(2) at a maximal specific rate of 0.4 U (micromol/min) per mg protein with an apparent K(m) for O(2) of 30 microM. The reaction was not inhibited by cyanide. The oxidase activity was traced back to a coenzyme F(420)-dependent enzyme that was purified to apparent homogeneity and that catalyzed the oxidation of 2 F(420)H(2) with 1 O(2) to 2 F(420) and 2 H(2)O. The apparent K(m) for F(420) was 30 microM and that for O(2) was 2 microM with a V(max) of 240 U/mg at 37 degrees C and pH 7.6, the pH optimum of the oxidase. The enzyme did not use NADH or NADPH as electron donor or H(2)O(2) as electron acceptor and was not inhibited by cyanide. The 45-kDa protein, whose gene was cloned and sequenced, contained 1 FMN per mol and harbored a binuclear iron center as indicated by the sequence motif H-X-E-X-D-X(62)-H-X(18)-D-X(60)-H. Sequence comparisons revealed that the F(420)H(2) oxidase from M. arboriphilus is phylogenetically closely related to FprA from Methanothermobacter marburgensis (71% sequence identity), a 45-kDa flavoprotein of hitherto unknown function, and to A-type flavoproteins from bacteria (30-40%), which all have dioxygen reductase activity. With heterologously produced FprA from M. marburgensis it is shown that this protein is also a highly efficient F(420)H(2) oxidase and that it contains 1 FMN and 2 iron atoms. The presence of F(420)H(2) oxidase in methanogenic archaea may explain why some methanogens, e.g., the Methanobrevibacter species in the termite hindgut, cannot only tolerate but thrive under microoxic conditions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs / genetics
  • Amino Acid Sequence
  • Cloning, Molecular
  • Conserved Sequence
  • Cyanides / pharmacology
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / isolation & purification
  • Enzyme Inhibitors / pharmacology
  • Enzyme Stability
  • Flavoproteins / genetics
  • Flavoproteins / physiology
  • Genes, Bacterial
  • Hydrogen / metabolism*
  • Hydrogen Peroxide / metabolism
  • Hydrogen-Ion Concentration
  • Methanobacteriaceae / enzymology
  • Methanobrevibacter / enzymology*
  • Molecular Weight
  • NAD / metabolism
  • Oxidation-Reduction
  • Oxidoreductases / genetics
  • Oxidoreductases / isolation & purification*
  • Oxidoreductases / metabolism*
  • Oxygen / metabolism*
  • Phylogeny
  • Riboflavin / analogs & derivatives*
  • Riboflavin / metabolism*
  • Sequence Alignment
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Temperature

Substances

  • Cyanides
  • DNA, Bacterial
  • Enzyme Inhibitors
  • Flavoproteins
  • NAD
  • coenzyme F420
  • Hydrogen
  • Hydrogen Peroxide
  • Oxidoreductases
  • Oxygen
  • Riboflavin