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Figure 4 | BMC Microbiology

Figure 4

From: The respiratory molybdo-selenoprotein formate dehydrogenases of Escherichia coli have hydrogen: benzyl viologen oxidoreductase activity

Figure 4

Analysis of H 2 - and formate-oxidizing activities of Fdh-N/O in different mutant backgrounds. Small-scale cultures of each strain were grown in TGYEP medium in the absence (A) or presence of nitrate (B). Extracts derived from the strains indicated were separated by non-denaturing PAGE and subsequently stained for H2: BV oxidoreductase (top panel), H2: PMS/NBT oxidoreductase (middle panel) or formate: PMS/NBT oxidoreductase (bottom panel) enzyme activity as described in the Methods section. Equivalent amounts of Triton X-100-treated crude extract (25 μg of protein) were applied to each lane. The activity band due to Fdh-N/Fdh-O is labelled by an arrow. The activity band due to hydrogenase 2 (Hyd-2) is also labelled in the top panel of part A and was used as a loading control for the experiment. Note the Hyd-2 activity can only be identified as a H2: BV oxidoreductase activity. The asterisk indicates hydrogenase activity associated with incompletely solubilised membrane material. The gel stained for H2: BV oxidoreductase activity was incubated for 8 h, while the gels stained with PMS/NBT were incubated for 1 h. In the interests of clarity, lanes were labelled based on the key genotype of the strain used. Lanes: MC4100 (wild type); FTD147 (ΔhyaB ΔhybC ΔhycE); FTD147 Δfnr signifies CP1104; ΔfdhE signifies JW3862 (ΔfdhE); ΔfdhE/pfdhE signifies JW3862 complemented with plasmid pCA24N-fdhE+; ΔfdhD signifies JW3866 (ΔfdhD); ΔfdhD/pfdhD signifies JW3866 complemented with plasmid pCA24N-fdhD+; ΔfdnG signifies JW1470 (ΔfdnG); ΔfdnG/pfdnG signifies JW1470 complemented with plasmid pCA24N-fdnG+; ΔfdoG signifies JW3865 (ΔfdoG); ΔfdoG/pfdoG signifies JW3865 complemented with plasmid pCA24N-fdoG+; ΔfdoG/pfdhE signifies JW3865 complemented with plasmid pCA24N-fdhE+. Note that BW25113 had an identical phenotype in these experiments to MC4100.

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