Molecular ecology and biochemistry of cyclohexenoyl-CoA hydrolysing/forming enzymes

在厌氧细菌中，芳香烃如BTEX通过中心中间体苯甲酰辅酶a降解。后者被所有芳香族化合物降解厌氧菌中存在的所谓苯甲酰辅酶a降解途径的酶进一步代谢。在之前的资助期内，我们开发并应用了一种基于功能pcr的检测组合，该组合专门用于编码苯甲酰辅酶a降解途径特征酶的保守基因。目标酶包括兼性厌氧菌（bcrC/ bzdn）和专性厌氧菌（bamb）的环切割水解酶（bamA-assay）、苯环去芳香化苯甲酰辅酶a还原酶（bcr）。这些检测方法在环境场所的应用已获批准。在下一个资助期内，我们将应用/修改mRNA检测和扩展应用（微阵列和/或扩增子测序）的分析。在第二个子项目中，我们研究了在巴豆酸盐和/或苯甲酸盐发酵过程中，在嗜酸养合菌中操作的苯甲酰辅酶a反向降解途径所涉及的酶。出乎意料的是，我们发现了一系列的二烯基辅酶a /单烯基辅酶a歧化反应参与了环己烯基辅酶a的形成。参与这些不寻常反应的酶将在下一个资助期内进行分离和表征。

In anaerobic bacteria aromatic hydrocarbons such as BTEX are degraded via the central intermediate benzoyl-CoA. The latter is further metabolized by enzymes of the so-called benzoyl-CoA degradation pathway present in all aromatic compound degrading anaerobic bacteria. In the preceding funding period we developed and applied a combination of functional PCR-based assays specific for conserved genes coding for characteristic enzymes of the benzoyl-CoA degradation pathway. The enzymes targeted comprised the ring cleaving hydrolase (bamA-assay), the benzene ring dearomatizing benzoyl-CoA reductases (BCRs) of facultative anaerobes (bcrC/bzdN-assays) and of obligate anaerobes (bamB-assay). The application of these assays at environmental sites was approved. In the next funding period we will apply/modify the assays for mRNA detection and extended applications (microarrays and/or amplicon sequencing). In a second subporoject we studied enzymes involved in a reverse benzoyl-CoA degradation pathway operating in Syntrophus aciditrophicus during crotonate and/or benzoate fermentation. Unexpectedly, we identified that a series of dienoyl-CoA/monoenoyl-CoA disproportionation reactions are involved in cyclohexanoyl-CoA formation. The enzymes involved in these unusual reactions will be isolated and characterized in the next funding period.