New aerobic benzoate oxidation pathway via benzoyl-coenzyme A and 3-hydroxybenzoyl-coenzyme A in a denitrifying Pseudomonas sp.
A denitrifying Pseudomonas sp. is able to oxidize aromatic compounds compounds completely to CO2, both aerobically and anaerobically. It is shown that benzoate is aerobically oxidized by a new degradation pathway via benzoyl-coenzyme A (CoA) and 3-hydroxybenzoyl-CoA. The organism grew aerobically with benzoate, 3-hydroxybenzoate, and gentisate; catechol, 2-hydroxybenzoate, and protocatechuate were not used, and 4-hydroxybenzoate was a poor substrate. Mutants were obtained which were not able to utilize benzoate as the sole carbon source aerobically but still used 3-hydroxybenzoate or gentisate. Simultaneous adaptation experiments with whole cells seemingly suggested a sequential induction of enzymes of a benzoate oxidation pathway via 3-hydroxybenzoate and gentisate. Cells grown aerobically with benzoate contained a benzoate-CoA ligase (AMP forming) (0.1 mumol min-1 mg-1) which converted benzoate but not 3-hydroxybenzoate into its CoA thioester. The enzyme of 130 kDa composed of two identical subunits of 56 kDa was purified and characterized. Cells grown aerobically with 3-hydroxybenzoate contained a similarly active CoA ligase for 3-hydroxybenzoate, 3-hydroxybenzoate-CoA ligase (AMP forming). Extracts from cells grown aerobically with benzoate catalyzed a benzoyl-CoA- and flavin adenine dinucleotide-dependent oxidation of NADPH with a specific activity of at least 25 nmol NADPH oxidized min-1 mg of protein-1; NADH and benzoate were not used. This new enzyme, benzoyl-CoA 3-monooxygenase, was specifically induced during aerobic growth with benzoate and converted [U-14C]benzoyl-CoA stoichiometrically to [14C]3-hydroxybenzoyl-CoA.
Altenschmidt
U
U
Abteilung Angewandte Mikrobiologie, University of Ulm, Germany.
Oswald
B
B
Steiner
E
E
Herrmann
H
H
Fuchs
G
G
D000214:Acyl Coenzyme A / Q000378:metabolism*; D000332:Aerobiosis / Q000502:physiology; D001565:Benzoates / Q000378:metabolism*; D019817:Benzoic Acid; D003066:Coenzyme A Ligases / Q000378:metabolism; D005841:Gentisates / Q000378:metabolism; D062385:Hydroxybenzoates / Q000378:metabolism; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009154:Mutation / Q000502:physiology; D010084:Oxidation-Reduction; D011549:Pseudomonas / Q000254:growth & development / Q000378:metabolism*; D013379:Substrate Specificity
C082323:3-hydroxybenzoyl-coenzyme A; D000214:Acyl Coenzyme A; D001565:Benzoates; D005841:Gentisates; D062385:Hydroxybenzoates; C032948:3-hydroxybenzoic acid; C037817:benzoyl-coenzyme A; D019817:Benzoic Acid; D006899:Mixed Function Oxygenases; C082322:benzoyl-CoA 3-monooxygenase; D003066:Coenzyme A Ligases; C055455:benzoate coenzyme A ligase