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A fluoride-insensitive inorganic pyrophosphatase isolated from Methanothrix soehngenii


An inorganic pyrophosphatase [E.C.] was isolated from Methanothrix soehngenii. In three steps the enzyme was purified 400-fold to apparent homogeneity. The molecular mass estimated by gelfiltration was 139±7 kDa. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis indicated that the enzyme is composed of subunits with molecular masses of 35 and 33 kDa in an α 2 β 2 oligomeric structure. The enzyme catalyzed the hydrolysis of inorganic pyrophosphate, tri-and tetrapolyphosphate, but no activity was observed with a variety of other phosphate esters. The cation Mg2+ was required for activity. The pH optimum was 8 at 1 mM PP i and 5 mM Mg2+. The enzyme was heat-stable, insensitive to molecular oxygen and not inhibited by fluoride. Analysis of the kinetic properties revealed an apparent K m for PP i of 0.1 mM in the presence of 5 mM Mg2+. The V max was 590 μmol of pyrophosphate hydrolyzed per min per mg protein, which corresponds to a K cat of 1400 per second.

The enzyme was found in the soluble enzyme fraction after ultracentrifugation, when cells were disrupted by French Press. Upto 5% of the pyrophosphatase was associated with the membrane fraction, when gentle lysis procedyre were applied.

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phenylmethylsulfonyl fluoride


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Jetten, M.S.M., Fluit, T.J., Stams, A.J.M. et al. A fluoride-insensitive inorganic pyrophosphatase isolated from Methanothrix soehngenii . Arch. Microbiol. 157, 284–289 (1992).

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Key words

  • Methanothrix soehngenii
  • Acetate degradation
  • Energetics
  • Inorganic pyrophosphatase
  • Fluoride inhibition