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. 1969 Sep;99(3):720–729. doi: 10.1128/jb.99.3.720-729.1969

Nitrate Reductase Complex of Escherichia coli K-12: Participation of Specific Formate Dehydrogenase and Cytochrome b1 Components in Nitrate Reduction

José Ruiz-Herrera a,1, J A DeMoss a
PMCID: PMC250087  PMID: 4905536

Abstract

The participation of distinct formate dehydrogenases and cytochrome components in nitrate reduction by Escherichia coli was studied. The formate dehydrogenase activity present in extracts prepared from nitrate-induced cells of strain HfrH was active with various electron acceptors, including methylene blue, phenazine methosulfate, and benzyl viologen. Certain mutants which are unable to reduce nitrate had low or undetectable levels of formate dehydrogenase activity assayed with methylene blue or phenazine methosulfate as electron acceptor. Of nine such mutants, five produced gas when grown anaerobically without nitrate and possessed a benzyl viologen-linked formate dehydrogenase activity, suggesting that distinct formate dehydrogenases participate in the nitrate reductase and formic hydrogenlyase systems. The other four mutants formed little gas when grown anaerobically in the absence of nitrate and lacked the benzyl viologen-linked formate dehydrogenase as well as the methylene blue or phenazine methosulfate-linked activity. The cytochrome b1 present in nitrate-induced cells was distinguished by its spectral properties and its genetic control from the major cytochrome b1 components of aerobic cells and of cells grown anaerobically in the absence of nitrate. The nitrate-specific cytochrome b1 was completely and rapidly reduced by 1 mm formate but was not reduced by 1 mm reduced nicotinamide adenine dinucleotide; ascorbate reduced only part of the cytochrome b1 which was reduced by formate. When nitrate was added, the formate-reduced cytochrome b1 was oxidized with biphasic kinetics, but the ascorbate-reduced cytochrome b1 was oxidized with monophasic kinetics. The inhibitory effects of n-heptyl hydroxyquinoline-N-oxide on the oxidation of cytochrome b1 by nitrate provided evidence that the nitrate-specific cytochrome is composed of two components which have different redox potentials but identical spectral properties. We conclude from these studies that nitrate reduction in E. coli is mediated by the sequential operation of a specific formate dehydrogenase, two specific cytochrome b1 components, and nitrate reductase.

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Selected References

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