FIG 3.
Activity assay for purified Hdr. (A) Proposed enzyme activity with either H2 or formate as the electron donor. (B) Time course of metronidazole reduction with protein purified from strains expressing hdrB-His. The assay was performed at 55°C in 0.7-ml anaerobic cuvettes containing 0.5 ml final assay volume with 80% H2 in the headspace. The final assay mixture consisted of 1.6 M potassium phosphate (pH 7), 180 μM metronidazole, 144 μM CoB-S-S-CoM, 3 μg protein, and 30 mM sodium formate. The H2-dependent assay was started by addition of CoB-S-S-CoM. The formate-dependent assay was started by addition of sodium formate. (C) Average specific activity of metronidazole (Mtz) reduction in the presence of H2 (0.23 μmol · min−1 mg−1) or in the presence of formate (4.52 μmol · min−1 mg−1). –, the background rate of Mtz reduction without CoB-S-S-CoM. Data are averages and standard deviations from four independent experiments.