Occurrence, genes and expression of the W/Se‐containing class II benzoyl‐coenzyme A reductases in anaerobic bacteria

Abstract

Benzoyl-coenzyme A (CoA) reductases (BCRs) are key enzymes in the anaerobic degradation of aromatic compounds and catalyse the reductive dearomatization of benzoyl-CoA to cyclohexa-1,5-dienoyl-1-carboxyl-CoA. Class I BCRs are ATP-dependent FeS enzymes, whereas class II BCRs are supposed to be ATP-independent and contain W, FeS clusters, and most probably selenocysteine. The active site components of a putative eight subunit class II BCR, BamBCDEFGHI, were recently characterized in Geobacter metallireducens. In this organism bamB was identified as structural gene for the W-containing active site subunit; bamF was predicted to code for a selenocysteine containing electron transfer subunit. In this work the occurrence and expression of BCRs in a number of anaerobic, aromatic compound degrading model microorganisms was investigated with a focus on the BamB and BamF components. Benzoate-induced class II BCR in vitro activities were determined in the soluble protein fraction in all obligately anaerobic bacteria tested. Where applicable, the results were in agreement with Western blot analysis using BamB targeting antibodies. By establishing a specific bamB targeting PCR assay, bamB homologues were identified in all tested obligately anaerobic bacteria with the capacity to degrade aromatic compounds; a number of bamB sequences from Gram-negative/positive sulfate-reducing bacteria were newly sequenced. In several organisms at least two bamB paralogues per genome were identified; however, in nearly all cases only one of them was transcribed during growth on an aromatic substrate. These benzoate-induced bamB genes are proposed to code for the active site subunit of class II BCRs; the major part of them group into a phylogenetic subcluster within the bamB homologues. Results from in silico analysis suggested that all class II BCRs contain selenocysteine in the BamF, and in many cases also in the BamE subunit. The results obtained indicate that the distribution of the two classes of BCRs in anaerobic bacteria appears to be strictly ruled by the available free energy from the oxidation of the aromatic carbon source rather than by phylogenetic relationships.

Cyclohexanecarboxyl-Coenzyme A (CoA) and Cyclohex-1-ene-1-Carboxyl-CoA Dehydrogenases, Two Enzymes Involved in the Fermentation of Benzoate and Crotonate in Syntrophus aciditrophicus

“…While ChCoA and Ch1CoA dehydrogenation could be measured in both directions with appropriate electron accepting/donating systems, no benzoyl-CoA reduction by Ch1CoA dehydrogenase was observed. In benzoate fermentation, BCoA reduction to Ch1,5CoA must then be catalyzed by a class II benzoyl-CoA reductase, which was recently identified in the obligate anaerobe Geobacter metallireducens (9,15) and predicted to be present in S. aciditrophicus (3,16). The results obtained in this work support the earlier hypothesis that formation of cyclohexane carboxylate from crotonate proceeds via CoA-ester intermediates of the reverse benzoyl-CoA degradation pathway (2).…”

Genome-scale analysis of anaerobic benzoate and phenol metabolism in the hyperthermophilic archaeon Ferroglobus placidus

“…The reaction catalyzed by this complex is reversible and ATP independent (Kung et al, 2009Lö ffler et al, 2010). It has been proposed that strict anaerobes must use class II BCRs because the amount of energy available from benzoate oxidation coupled to the reduction of Fe(III), sulfate or protons is not sufficient to support the substantial energetic requirement of the ATP-dependent class I BCR reaction (Schöcke and Schink, 1999;Kung et al, 2009Kung et al, , 2010Lö ffler et al, 2010).…”

Enzymes Involved in a Novel Anaerobic Cyclohexane Carboxylic Acid Degradation Pathway

“…However, it differs from the Thauera aromatica type of benzoyl-CoA degradation pathway present in all other known aromatic compound-degrading anaerobic bacteria. In denitrifying, sulfate-reducing, Fe(III)-reducing, and fermenting bacteria, the class I and II benzoyl-CoA reductases reduce their substrate only by two electrons, yielding cyclohex-1,5-diene-1-carboxyl-CoA (CHdieneCoA) (18,19). As a consequence, a different set of enzymes is involved in CHdieneCoA degradation, including specific hydratases, dehydrogenases, and ring-cleaving hydrolases that form 3-hydroxypimeloyl-CoA and not pimeloyl-CoA.…”