Enzymes of the benzoyl-coenzyme A degradation pathway in the hyperthermophilic archaeon Ferroglobus placidus
Enzymes of the benzoyl-coenzyme A degradation pathway in the hyperthermophilic archaeon Ferroglobus placidus
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DOI：
10.1111/1462-2920.12785
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发表时间：
2015-09-01
影响因子：
5.1
通讯作者：
Boll, Matthias
中科院分区：
文献类型：
--
作者：
Schmid, Georg;Rene, Sandra Bosch;Boll, Matthias
呼吸Fe(III)的高地铁球藻是已知的唯一一种芳香底物完全降解为CO2的古细菌和嗜热菌。最近的基因组和转录组分析提出了一个类似于沼泽红假单胞菌中发现的苯甲酰辅酶A(CoA)的降解途径，其中包括形成依赖于ATP的关键酶苯甲酰辅酶A还原酶(BCR)的环己烯-1-羧基-辅酶A(1-Enoyl-CoA)。在这项工作中，我们首先通过体外研究证明，苯甲酰辅酶A依赖于ATP被两个电子还原为环己烷-1，5-二烯-辅酶A(1，5-二烯-辅酶A)，后者通过水合进一步降解为6-羟基环己基-1-烯-1-羧基-辅酶A(6-OH-1-Enoyl-CoA)；当加入NAD(+)时，后者随后被转化为氧化中间体。BCR的四个候选基因异源表达，富含氧敏感的酶催化苯甲酰辅酶A的两电子还原为1，5-二烯酰辅酶A。一个先前被指定为2，3-二脱氢吡喃甲酰-辅酶A水合酶的基因被异源表达，并被证明是典型的不接受1-烯基-辅酶A的1，5-二烯酰辅酶A水合酶。异源表达了一个先前属于1-烯醇基-辅酶A水合酶的基因，并鉴定其编码一个双功能巴豆酶/3-羟基-丁酰辅酶A脱氢酶。总之，这些结果一致地提供了生化证据，表明Placidus和可能其他古生菌主要通过Thauera/Azoarcus类型降解芳香物，而不是或仅在很小程度上通过预测的R.palustris类型的苯甲酰-CoA降解途径。
The Fe(III)-respiring Ferroglobus placidus is the only known archaeon and hyperthermophile for which a complete degradation of aromatic substrates to CO2 has been reported. Recent genome and transcriptome analyses proposed a benzoyl-coenzyme A (CoA) degradation pathway similar to that found in the phototrophic Rhodopseudomonas palustris, which involves a cyclohex-1-ene-1-carboxyl-CoA (1-enoyl-CoA) forming, ATP-dependent key enzyme benzoyl-CoA reductase (BCR). In this work, we demonstrate, by first in vitro studies, that benzoyl-CoA is ATP-dependently reduced by two electrons to cyclohexa-1,5-dienoyl-CoA (1,5-dienoyl-CoA), which is further degraded by hydration to 6-hydroxycyclohex-1-ene-1-carboxyl-CoA (6-OH-1-enoyl-CoA); upon addition of NAD(+), the latter was subsequently converted to -oxidation intermediates. The four candidate genes of BCR were heterologously expressed, and the enriched, oxygen-sensitive enzyme catalysed the two-electron reduction of benzoyl-CoA to 1,5-dienoyl-CoA. A gene previously assigned to a 2,3-didehydropimeloyl-CoA hydratase was heterologously expressed and shown to act as a typical 1,5-dienoyl-CoA hydratase that does not accept 1-enoyl-CoA. A gene previously assigned to a 1-enoyl-CoA hydratase was heterologously expressed and identified to code for a bifunctional crotonase/3-OH-butyryl-CoA dehydrogenase. In summary, the results consistently provide biochemical evidence that F.placidus and probably other archaea predominantly degrade aromatics via the Thauera/Azoarcus type and not or only to a minor extent via the predicted R.palustris-type benzoyl-CoA degradation pathway.