CpdA is involved in amino acid metabolism in <i>Shewanella oneidensis</i> MR-1

<p>Cyclic 3′,5′-adenosine monophosphate (cAMP) phosphodiesterase (CPD) is an enzyme that catalyzes the hydrolysis of cAMP, a signaling molecule affecting diverse cellular and metabolic processes in bacteria. Some CPDs are also known to function in cAMP-independent manners, while their physiological roles remain largely unknown. Here, we investigated physiological roles of CPD in <i>Shewanella oneidensis</i> MR-1, a model environmental bacterium, and report that CPD is involved in amino-acid metabolism. We found that a CPD-deficient mutant of MR-1 (Δ<i>cpdA</i>) showed decreased expression of genes for the synthesis of methionine, <i>S</i>-adenosylmethionine, and histidine and required these three compounds to grow in minimal media. Interestingly, deletion of adenylate cyclases in Δ<i>cpdA</i> did not restore the ability to grow in minimal media, indicating that the amino acid requirements were not due to the accumulation of cAMP. These results suggest that CPD is involved in the regulation of amino acid metabolism in MR-1 in a cAMP-independent manner.</p> <p>cAMP and cGMP levels and growth rates of cells aerobically grown in lactate minimal medium are shown in panels A, B, and C, respectively.</p

Polarization (open squares) and power (closed squares) curves for the methanol-fed MFC.

<p>Polarization (open squares) and power (closed squares) curves for the methanol-fed MFC.</p

Comparison of the total lengths of large contigs affiliated with different phyla as determined by MEGAN or BLSOM analyses of the metagenome data.

<p>Comparison of the total lengths of large contigs affiliated with different phyla as determined by MEGAN or BLSOM analyses of the metagenome data.</p

Catabolic pathway for methanol/acetate conversion in the methanol-fed MFC predicted from the metagenome data (A), and phylum-level distributions of genes assigned to each catabolic step (B).

<p>Step I, methanol:THF methyltransferase; II, acetyl-CoA synthase (EC.2.3.1.169); III, carbon monoxide dehydrogenase (EC.1.2.7.4); IV, acetyl-CoA synthetase (EC.6.2.1.1); V, phosphate acetyltransferase (EC.2.3.1.8); and VI, acetate kinase (EC.2.7.2.1).</p

Typical time courses of cell voltage (A), methanol concentration (B), and acetate concentration (C), after supplementation of the MFC with 10 mM methanol.

<p>In panels B and C, data are means ± SD (n = 3), and error bars are shown when they are larger than symbols.</p

Summary of numerical data for the metagenomic analyses of microbes associated with the anode biofilm in the methanol-fed MFC.

<p>Summary of numerical data for the metagenomic analyses of microbes associated with the anode biofilm in the methanol-fed MFC.</p

Distribution of microbes in the methanol-fed MFC.

<p>(A) Total protein contents showing amounts of microbes associated with the anode biofilm, cathode biofilm, and electrolyte. (B) Results of an anode-exchange experiment, in which cell voltages of methanol-fed MFCs were measured after the microbe-bearing anode was transferred to a reactor containing fresh electrolyte (black line) and a new anode was placed in the spent electrolyte of the initial reactor (gray line).</p

Metagenomic Analyses Reveal the Involvement of Syntrophic Consortia in Methanol/Electricity Conversion in Microbial Fuel Cells

<div><p>Methanol is widely used in industrial processes, and as such, is discharged in large quantities in wastewater. Microbial fuel cells (MFCs) have the potential to recover electric energy from organic pollutants in wastewater; however, the use of MFCs to generate electricity from methanol has not been reported. In the present study, we developed single-chamber MFCs that generated electricity from methanol at the maximum power density of 220 mW m⁻² (based on the projected area of the anode). In order to reveal how microbes generate electricity from methanol, pyrosequencing of 16S rRNA-gene amplicons and Illumina shotgun sequencing of metagenome were conducted. The pyrosequencing detected in abundance <i>Dysgonomonas</i>, <i>Sporomusa</i>, and <i>Desulfovibrio</i> in the electrolyte and anode and cathode biofilms, while <i>Geobacter</i> was detected only in the anode biofilm. Based on known physiological properties of these bacteria, it is considered that <i>Sporomusa</i> converts methanol into acetate, which is then utilized by <i>Geobacter</i> to generate electricity. This speculation is supported by results of shotgun metagenomics of the anode-biofilm microbes, which reconstructed relevant catabolic pathways in these bacteria. These results suggest that methanol is anaerobically catabolized by syntrophic bacterial consortia with electrodes as electron acceptors.</p></div

Gene clusters containing putative methanol:THF methyltransferases (black arrows) in metagenome contig NODE_348 and the genome of <i>Sporomusa ovata</i>.

<p>Possible genes encoding transcriptional regulators for methyltransferases are indicated with gray arrows. Results of BLAST search for the genes are described in the table.</p

Potential dependency of the microbial current.

<p>(A) Microbial current by wild-type cells after 1 h (triangles), 5 h (circles), and 20 h (squares) of electrochemical cultivation at the indicated potentials. (B) Microbial current by wild-type cells just before (black squares) and after (white squares) the addition of malonic acid into the electrochemical system. (C) Microbial current by <i>sdh</i> deficient mutant cells after 20 h of electrochemical cultivation.</p