Unusual enzyme complexes in anaerobic aromatic metabolism of Geobacter metallireducens

同时性厌氧细菌的厌氧芳香代谢已被广泛研究。由于整体能量代谢的限制，许多关键的酶促过程在专性厌氧细菌中有所不同。例如，在专性厌氧菌中，依赖于atp的去芳香化I类苯甲酰辅酶a还原酶（bcrabcd -亚基，FeS-cofactors）被atp独立的II类BCR （bambcdefgi -亚基，W/FeS/SeCys/FAD-cofactors）所取代。在上一个资助期，我们从Geobacter metallireducens中分离并表征了以前未知的含有活性位点亚基BamBC的W/ fes簇，BamBC的晶体结构使我们能够首次了解其机制。G. metallireducens中另一种不寻常的酶复合物是对甲酚甲基羟化酶（PCMH， pcmgij亚单位），它催化厌氧对甲酚代谢的第一步。它与其他pcmh的不同之处在于其在膜上的紧密位置，可能与细胞色素bc1样复合体（pcmcdef亚基）有关，并且其不寻常的亚基/辅因子组成。在上一次资助期间，我们分离并表征了可溶性PmcGIJ成分。在下一个资助期内，II类BCR和PCMH的整个酶复合物（包括迄今为止缺失的成分）将被分离出来，并通过动力学、光谱和结构方法进行研究。主要目的是深入了解新型多组分酶复合物在专性厌氧细菌中不寻常的能量转导/保存过程中的功能。

The anaerobic aromatic metabolism has extensively been studied in facultatively anaerobic bacteria. Due to limitations in the overall energy metabolism many key enzymatic processes differ in obligately anaerobic bacteria. E.g., the ATP-dependent, dearomatizing class I benzoyl-CoA reductase (BcrABCD-subunits, FeS-cofactors) is replaced by an ATP-independent class II BCR in obligate anaerobes (BamBCDEFGHI-subunits, W/FeS/SeCys/FAD-cofactors). In the last funding period we have isolated and characterized the previsouly unknown W/FeS-cluster containing active site subunits BamBC from Geobacter metallireducens, and the crystal structure of BamBC enabled first insights into the mechanism. Another unusual enzyme complex in G. metallireducens is p-cresol methylhydroxylase (PCMH, PcmGIJ-subunits) catalyzing the first steps in anaerobic p-cresol metabolism. It differs from other PCMHs by its tight location in the membrane, possibly in association with a cytochrome bc1-like complex (PcmCDEF-subunits) and by its unusual subunit/cofactor-composition. In the last funding period we have isolated and characterized the soluble PmcGIJ components. In the next funding period both entire enzyme complexes of the class II BCR and PCMH including the so far missing components will be isolated and studied by kinetic, spectroscopic and structural methodologies. A main objective is to obtain insights into the function of novel multi component enzyme complexes involved in unusual energy transduction/conservation processes in obligately anaerobic bacteria.