Institute of Biochemistry, University of Leipzig, Leipzig, Germany Faculty of Biology and Environmental Protection, University of Silesia, Katowice, Poland.
Environ Microbiol. 2011 Mar;13(3):696-709. doi: 10.1111/j.1462-2920.2010.02374.x. Epub 2010 Nov 18.
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.
苯甲酰辅酶 A（CoA）还原酶（BCRs）是芳香族化合物厌氧降解的关键酶，催化苯甲酰-CoA 的还原去芳构化为环己-1,5-二烯酰-1-羧基-CoA。I 类 BCR 是依赖 ATP 的 FeS 酶，而 II 类 BCR 则被认为是不依赖 ATP 的，包含 W、FeS 簇，并且很可能包含硒代半胱氨酸。最近在 Geobacter metallireducens 中对假定的八亚基 II 类 BCR（BamBCDEFGHI）的活性位点成分进行了表征。在该生物中，bamB 被鉴定为含有 W 的活性位点亚基的结构基因；bamF 被预测为编码含有硒代半胱氨酸的电子转移亚基。在这项工作中，研究了一些厌氧、芳香族化合物降解模式微生物中 BCR 的发生和表达，重点关注 BamB 和 BamF 成分。在所有测试的严格厌氧菌的可溶性蛋白部分中，均测定了苯甲酸盐诱导的体外 II 类 BCR 活性。在适用的情况下，结果与使用 BamB 靶向抗体的 Western blot 分析一致。通过建立特定的 BamB 靶向 PCR 测定法，在所有具有芳香族化合物降解能力的严格厌氧菌中鉴定出 BamB 同源物；从革兰氏阴性/阳性硫酸盐还原菌中获得了一些 BamB 序列的新测序结果。在几种生物中，每个基因组至少鉴定出两个 BamB 旁系同源物；然而，在几乎所有情况下，只有一个在以芳香族底物生长时被转录。这些苯甲酸盐诱导的 BamB 基因被提议编码 II 类 BCR 的活性位点亚基；它们中的大部分在 BamB 同源物中形成一个系统发育亚群。计算机分析结果表明，所有 II 类 BCR 都在 BamF 中包含硒代半胱氨酸，并且在许多情况下也在 BamE 亚基中包含硒代半胱氨酸。所得结果表明，厌氧细菌中两类 BCR 的分布似乎严格由芳香族碳源氧化提供的可用自由能决定，而不是由系统发育关系决定。
