Purification of glutaryl-CoA dehydrogenase from Pseudomonas sp., an enzyme involved in the anaerobic degradation of benzoate.
Cell-free extracts of Pseudomonas sp. strains KB 740 and K 172 both contained high levels of glutaryl-CoA dehydrogenase when grown anaerobically on benzoate or other aromatic compounds and with nitrate as electron acceptor. These aromatic compounds have in common benzoyl-CoA as the central aromatic intermediate of anaerobic metabolism. The enzymatic activity was almost absent in cells grown aerobically on benzoate regardless whether nitrate was present. Glutaryl-CoA dehydrogenase activity was also detected in cell-free extracts of Rhodopseudomonas, Rhodomicrobium and Rhodocyclus after phototrophic growth on benzoate. Parallel to the induction of glutaryl-CoA dehydrogenase as measured with ferricenium ion as electron acceptor, an about equally high glutaconyl-CoA decarboxylase activity was detected in cell-free extracts. The latter activity was measured with the NAD-dependent assay, as described for the biotin-containing sodium ion pump glutaconyl-CoA decarboxylase from glutamate fermenting bacteria. Glutaryl-CoA dehydrogenase was purified to homogeneity from both Pseudomonas strains. The enzymes catalyse the decarboxylation of glutaconyl-CoA at about the same rate as the oxidative decarboxylation of glutaryl-CoA. The green enzymes are homotetramers (m = 170 kDa) and contain 1 mol FAD per subunit. No inhibition was observed with avidin indicating the absence of biotin. The N-terminal sequences of the enzymes from both strains are similar (65%).
Härtel
U
U
Laboratorium für Mikrobiologie, Fachbereich Biologie, Philipps-Universität, Marburg, Federal Republic of Germany.
Eckel
E
E
Koch
J
J
Fuchs
G
G
Linder
D
D
Buckel
W
W
D000595:Amino Acid Sequence; D001565:Benzoates / Q000378:metabolism*; D019817:Benzoic Acid; D001673:Biodegradation, Environmental; D002262:Carboxy-Lyases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D050770:Glutaryl-CoA Dehydrogenase; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D044925:Oxidoreductases Acting on CH-CH Group Donors; D010100:Oxygen; D011549:Pseudomonas / Q000145:classification / Q000201:enzymology*
D001565:Benzoates; D019817:Benzoic Acid; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; D050770:Glutaryl-CoA Dehydrogenase; D002262:Carboxy-Lyases; C023060:glutaconyl CoA decarboxylase; D010100:Oxygen