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Deletion of genes encoding cytochrome oxidases and quinol monooxygenase blocks the aerobic-anaerobic shift in Escherichia coli K-12 MG1655.

ABSTRACT: The constitutive activation of the anoxic redox control transcriptional regulator (ArcA) in Escherichia coli during aerobic growth, with the consequent production of a strain that exhibits anaerobic physiology even in the presence of air, is reported in this work. Removal of three terminal cytochrome oxidase genes (cydAB, cyoABCD, and cbdAB) and a quinol monooxygenase gene (ygiN) from the E. coli K-12 MG1655 genome resulted in the activation of ArcA aerobically. These mutations resulted in reduction of the oxygen uptake rate by nearly 98% and production of d-lactate as a sole by-product under oxic and anoxic conditions. The knockout strain exhibited nearly identical physiological behaviors under both conditions, suggesting that the mutations resulted in significant metabolic and regulatory perturbations. In order to fully understand the physiology of this mutant and to identify underlying metabolic and regulatory reasons that prevent the transition from an aerobic to an anaerobic phenotype, we utilized whole-genome transcriptome analysis, (13)C tracing experiments, and physiological characterization. Our analysis showed that the deletions resulted in the activation of anaerobic respiration under oxic conditions and a consequential shift in the content of the quinone pool from ubiquinones to menaquinones. An increase in menaquinone concentration resulted in the activation of ArcA. The activation of the ArcB/ArcA regulatory system led to a major shift in the metabolic flux distribution through the central metabolism of the mutant strain. Flux analysis indicated that the mutant strain had undetectable fluxes around the tricarboxylic acid (TCA) cycle and elevated flux through glycolysis and anaplerotic input to oxaloacetate. Flux and transcriptomics data were highly correlated and showed similar patterns.

SUBMITTER: Portnoy VA

PROVIDER: S-EPMC2950451 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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Deletion of genes encoding cytochrome oxidases and quinol monooxygenase blocks the aerobic-anaerobic shift in Escherichia coli K-12 MG1655.

Portnoy Vasiliy A VA Scott David A DA Lewis Nathan E NE Tarasova Yekaterina Y Osterman Andrei L AL Palsson Bernhard Ø BØ

Applied and environmental microbiology 20100813 19

The constitutive activation of the anoxic redox control transcriptional regulator (ArcA) in Escherichia coli during aerobic growth, with the consequent production of a strain that exhibits anaerobic physiology even in the presence of air, is reported in this work. Removal of three terminal cytochrome oxidase genes (cydAB, cyoABCD, and cbdAB) and a quinol monooxygenase gene (ygiN) from the E. coli K-12 MG1655 genome resulted in the activation of ArcA aerobically. These mutations resulted in reduc ...[more]

PMID: 20709841

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Project description:To exchange electrons with extracellular substrates, some microorganisms employ extracellular electron transfer (EET) pathways that physically connect extracellular redox reactions to intracellular metabolic activity. These pathways are made of redox and structural proteins that work cooperatively to transfer electrons between extracellular substrates and the cytoplasmic membrane. Crucial to the bacterial and archaeal EET pathways are the quinol oxidases and/or quinone reductases in the cytoplasmic membrane where they recycle the quinone/quinol pool in the cytoplasmic membrane during EET reaction. Up to date, three different families of quinol oxidases and/or quinone reductases involved in bacterial EET have been discovered. They are the CymA, CbcL/MtrH/MtoC, and ImcH families of quinol oxidases and/or quinone reductases that are all multiheme c-type cytochromes (c-Cyts). To investigate to what extent they are distributed among microorganisms, we search the bacterial as well as archaeal genomes for the homologs of these c-Cyts. Search results reveal that the homologs of these c-Cyts are only found in the Domain Bacteria. Moreover, the CymA homologs are only found in the phylum of Proteobacteria and most of them are in the Shewanella genus. In addition to Shewanella sp., CymA homologs are also found in other Fe(III)-reducing bacteria, such as of Vibrio parahaemolyticus. In contrast to CymA, CbcL/MtrH/MtoC, and ImcH homologs are much more widespread. CbcL/MtrH/MtoC homologs are found in 15 phyla, while ImcH homologs are found in 12 phyla. Furthermore, the heme-binding motifs of CbcL/MtrH/MtoC and ImcH homologs vary greatly, ranging from 3 to 23 and 6 to 10 heme-binding motifs for CbcL/MtrH/MtoC and ImcH homologs, respectively. Moreover, CymA and CbcL/MtrH/MtoC homologs are found in both Fe(III)-reducing and Fe(II)-oxidizing bacteria, suggesting that these families of c-Cyts catalyze both quinol-oxidizing and quinone-reducing reactions. ImcH homologs are only found in the Fe(III)-reducing bacteria, implying that they are only the quinol oxidases. Finally, some bacteria have the homologs of two different families of c-Cyts, which may improve the bacterial capability to exchange electrons with extracellular substrates.

Dataset Information

Deletion of genes encoding cytochrome oxidases and quinol monooxygenase blocks the aerobic-anaerobic shift in Escherichia coli K-12 MG1655.

Publications

Deletion of genes encoding cytochrome oxidases and quinol monooxygenase blocks the aerobic-anaerobic shift in Escherichia coli K-12 MG1655.

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