Zaar, AnnetteAnnetteZaarGescher, JohannesJohannesGescherEisenreich, WolfgangWolfgangEisenreichBacher, AdelbertAdelbertBacherFuchs, GeorgGeorgFuchs2023-02-102023-02-102004-08-11Molecular Microbiology 54 (1): 223-238 (2004-10-01)http://hdl.handle.net/11420/14784A new principle of aerobic aromatic metabolism has been postulated, which is in contrast to the known pathways. In various bacteria the aromatic substrate benzoate is first converted to its coenzyme A (CoA) thioester, benzoyl-CoA, which is subsequently attacked by an oxygenase, followed by a non-oxygenolytic fission of the ring. We provide evidence for this hypothesis and show that benzoyl-CoA conversion in the bacterium Azoarcus evansii requires NADPH, O 2 and two protein components, BoxA and BoxB. BoxA is a homodimeric 46 kDa iron-sulphur-flavoprotein, which acts as reductase. In the absence of BoxB, BoxA catalyses the benzoyl-CoA stimulated artificial transfer of electrons from NADPH to O2 via free FADH2 to produce H2O 2. Physiologically, BoxA uses NADPH to reduce BoxB, a monomeric 55 kDa iron-protein that acts as benzoyl-CoA oxygenase. The product of benzoyl-CoA oxidation was identified by NMR spectroscopy as its dihydrodiol derivative, 2,3-dihydro-2,3-dihydroxybenzoyl-CoA. This suggests that BoxBA act as a benzoyl-CoA dioxygenase/reductase. Unexpectedly, benzoyl-CoA transformation by BoxBA was greatly stimulated when another enoyl-CoA hydratase/isomerase-like protein, BoxC, was added that catalysed the further transformation of the dihydrodiol product formed from benzoyl-CoA. The benzoyl-CoA oxygenase system has very low similarity to known (di)oxygenase systems and is the first member of a new enzyme family.en1365-295820041223238Wiley-BlackwellBiowissenschaften, BiologieJournal Article10.1111/j.1365-2958.2004.04263.x15458418Other