Biochemistry of bacterial dissimilatory metal reduction

异化金属还原是一种呼吸过程，在各种细菌和微生物中起作用。这些微生物能够使用氧化金属如Fe（III）、Mn（IV）、Cr（VI）或U（VI）作为其厌氧电子传递链的末端电子受体。希瓦氏菌是研究这一过程的两种模式生物之一。然而，即使经过几十年的研究异化金属还原，电子传递到金属电子受体的必要的最小蛋白质集仍然是未知的。我正在寻求一种基于合成生物学的新方法来解决这个问题。因此，本研究的目标是通过表达大肠杆菌中潜在的关键基因，将大肠杆菌转化为异化金属还原剂。oneidensis。使用这种技术，我们已经建立了一个菌株，能够呼吸螯合铁（III）作为终端电子受体，从而显示了该方法的适用性。通过进一步插入来自S的电子传递链的可能化合物的基因。我们希望进入下一个层次，建立电子转移到晶体铁和锰的形式。我们将使用几种平行的方法检测还原链的潜在关键参与者，包括质谱，纯化和后续表征以及靶向基因缺失。已经进行了一系列关于还原链到三价铁的原理实验的证明。这些体外实验揭示了蛋白质间电子转移反应，并且也得到了体内实验的验证。设计的E.大肠杆菌菌株被认为进一步用于解决蛋白定位和复合物形成的问题，并作为进一步结晶研究的靶的异源表达的平台。

Dissimilatory metal reduction is a respiratory process that operates in various Bacteria and Archaea. These microorganisms are able to use oxidized metals like Fe(III), Mn(IV), Cr(VI), or U(VI) as terminal electron acceptors of their anaerobic electron transport chains. Shewanella oneidensis is one of the two model organisms to study this process. However, even after decades of research on dissimilatory metal reduction, the essential minimal protein set for the electron transport to metallic electron acceptors is still unknown. I am pursuing a novel synthetic biology based approach to tackle this problem. Therefore, the goal of the proposed study is to convert Escherichia coli into a dissimilatory metal reducer through expression of potential key genes from S. oneidensis. Using this technique we have already established a strain that is able to respire with chelated Fe(III) as terminal electron acceptor and have thereby shown the applicability of the method. Via the insertion of further genes for possible compounds of the electron transport chain from S. oneidensis we want to proceed to the next level, being to establish an electron transfer to crystalline iron and manganese forms. We will detect potential key players for the reduction chain to ferric iron and manganese using several parallel approaches, which include mass spectrometry, purification and following characterization as well as targeted gene deletions. A series of proof of principle experiments for the reduction chain to ferric iron were already conducted. These in vitro experiments revealed interprotein electron transfer reactions and were also verified by in vivo experiments. The designed E. coli strains are supposed to be further used to address questions of protein localization and complex formation and as a platform for heterologues expression of targets for further crystallization studies.