Structure and function of benzoyl-coenzyme A reductases

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The benzene ring represents the second most abundant biologically produced structure on earth (25% at terrestrial sites), and is present in lignin, fossil oil and many environmentally relevant aromatic compounds (e.g., solvents, pesticides, insecticides). Under anaerobic conditions, e.g. at marine/freshwater sediments or contaminated aquifers only anaerobic prokaryotes are capable of fully degrading aromatic compounds. In these organisms the multitude of monocyclic aromatic compounds are converted to the central intermediate benzoyl-coenzyme A that is dearomatized to a dienoyl-CoA by reductases. This reduction proceeds at the negative redox potential limit in biology and requires a coupling to an exergonic process. There are two fundamentally different biological solutions for enzymatic benzoyl-CoA reduction, catalyzed by the class I and II benzoyl-CoA reductases (BCRs). Class I BCRs are present in facultative anaerobes; they are composed of four BcrABCD subunits and contain three [4Fe-4S] clusters. Class I BCRs couple the endergonic electron transfer from reduced ferredoxin to the aromatic ring to a stoichiometric ATP-hydrolysis. The highly complex 1 MDa class II BCRs are composed of the eight BamBCDEFGHI subunits and contain W, Zn, FeS clusters, Se and FAD. They are proposed to drive benzoyl-CoA reduction by a flavin-based electron transfer process. The project aims understand the structure and function of enzymes catalyzing a reaction that in organic chemical synthesis requires extremely harsh conditions (Birch reduction using alkali metals as reductants). Numerous spectroscopic, kinetic, and structural studies will be carried out with class I and II BCRs from different organisms, and with molecular variants generated by the use of newly developed expression/manipulation platforms. The project synergistically connects the 20 year experience of the applicant with enzymes involved in anaerobic aromatic metabolism with state-of-the-art structural and spectroscopic methodologies provided by numerous national/international collaboration partners.