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Purification and properties of the H2-oxidizing (uptake) hydrogenase of the N2-fixing anaerobeClostridium pasteurianumW5

Authors
Journal
Biochemical and Biophysical Research Communications
0006-291X
Publisher
Elsevier
Publication Date
Volume
122
Issue
1
Identifiers
DOI: 10.1016/0006-291x(84)90431-5
Disciplines
  • Biology

Abstract

Abstract Clostridium pasteurianum has two distinct hydrogenases, the bidirectional hydrogenase and the H 2-oxidizing (uptake) hydrogenase. The H 2-oxidizing hydrogenase has been purified (up to 970-fold) to a specific activity of 17,600 μmol H 2 oxidized/min·mg protein (5 mM methylene blue) or 3.5 μmol H 2 produced/min·mg protein (1 mM methyl viologen). The uptake hydrogenase has a M r of 53,000 (one polypeptide chain). Depending upon how protein was measured, the Fe and S = contents (gatom/mol) were 4.7 and 5.2 (by the dye-binding assay) or 7.2 and 8.0 (by the Lowry method). Both reduced and oxidized forms of the enzyme gave electron paramagnetic resonance signals. The activation energy for H 2-production and H 2-oxidation by the uptake hydrogenase was 59.1 and 31.2 kJ/mol, respectively. In the exponential phase of growth, the ratio of uptake hydrogenase/bidirectional hydrogenase in NH 3-grown cells was much lower than that in N 2-fixing cells.

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