Methyl viologen hydrogenase II, a new member of the hydrogenase family from Methanobacterium thermoautotrophicum ΔH

G. J. Woo, A. Wasserfallen, R. S. Wolfe

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15 Citations (Scopus)

Abstract

Two methyl viologen hydrogenase (MVH) enzymes from Methanobacterium thermoautotrophicum ΔH have been separated (resolution, R(s) at 1.0) on a Mono Q column after chromatography on DEAE-Sephacel and Superose 6 Prep Grade. The newly discovered MVH (MVH II) was eluted at 0.5 M NaCl with a linear gradient of 0.45 to 0.65 M NaCl (100 ml). The previously described MVH (MVH I) eluted in a NaCl gradient at 0.56 M. The specific activities of MVH I and MVH II were 184.8 and 61.3 U/mg of protein, respectively, when enzyme activity was compared at pH 7.5, the optimal pH for MVH II. Gel electrophoresis in nondenaturing systems indicated that MVH I and MVH II had a similar molecular mass of 145 kDa. Denatured MVH II showed four protein bands (α, 50 kDa; β, 44 kDa; γ, 36 kDa; δ, 15 kDa), similar to MVH I. The N-terminal amino acid sequences of the α, γ, and δ subunits of MVH II were identical with the sequences of the equivalent subunits of MVH I. However, the N-terminal amino acid sequence of the β subunit of MVH II was totally different from the sequence of the β subunit of MVH I. Both MVH I and MVH II had the same optimal temperature of 60°C for maximum activity. The pH optima of MVH I and MVH II were 9.0 and 7.5, respectively. Most of the divalent metal ions tested significantly inhibited MVH I activity, but MVH II activity was only partially inhibited by some divalent cations. Both hydrogenases were shown to be stable for over 8 days at -20°C under anaerobic conditions. When exposed to air, 90% of MVH I activity was lost within 2 min; however, MVH II lost only 50% of its activity in 3 h.

Original languageEnglish
Pages (from-to)5970-5977
Number of pages8
JournalJournal of Bacteriology
Volume175
Issue number18
Publication statusPublished - 1993 Jan 1

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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