title: Phenol hydroxylase and toluene/o-xylene monooxygenase from Pseudomonas stutzeri OX1: interplay between two enzymes.
abstract: Degradation of aromatic hydrocarbons by aerobic bacteria is generally divided into an upper pathway, which produces dihydroxylated aromatic intermediates by the action of monooxygenases, and a lower pathway, which processes these intermediates down to molecules that enter the citric acid cycle. Bacterial multicomponent monooxygenases (BMMs) are a family of enzymes divided into six distinct groups. Most bacterial genomes code for only one BMM, but a few cases (3 out of 31) of genomes coding for more than a single monooxygenase have been found. One such case is the genome of Pseudomonas stutzeri OX1, in which two different monooxygenases have been found, phenol hydroxylase (PH) and toluene/o-xylene monooxygenase (ToMO). We have already demonstrated that ToMO is an oligomeric protein whose subunits transfer electrons from NADH to oxygen, which is eventually incorporated into the aromatic substrate. However, no molecular data are available on the structure and on the mechanism of action of PH. To understand the metabolic significance of the association of two similar enzymatic activities in the same microorganism, we expressed and characterized this novel phenol hydroxylase. Our data indicate that the PH P component of PH transfers electrons from NADH to a subcomplex endowed with hydroxylase activity. Moreover, a regulatory function can be suggested for subunit PH M. Data on the specificity and the kinetic constants of ToMO and PH strongly support the hypothesis that coupling between the two enzymatic systems optimizes the use of nonhydroxylated aromatic molecules by the draining effect of PH on the product(s) of oxidation catalyzed by ToMO, thus avoiding phenol accumulation.
authors.affiliation: Dipartimento di Chimica Biologica, Università di Napoli Federico II, 16-80134 Naples, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: Cafaro
authors.forename: Viviana
authors.initials: V
authors.lastname: Izzo
authors.forename: Roberta
authors.initials: R
authors.lastname: Scognamiglio
authors.forename: Eugenio
authors.initials: E
authors.lastname: Notomista
authors.forename: Paola
authors.initials: P
authors.lastname: Capasso
authors.forename: Annarita
authors.initials: A
authors.lastname: Casbarra
authors.forename: Piero
authors.initials: P
authors.lastname: Pucci
authors.forename: Alberto
authors.initials: A
authors.lastname: Di Donato
mesh_terms: D001483:Base Sequence; D001673:Biodegradation, Environmental; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D006841:Hydrocarbons, Aromatic / Q000378:metabolism; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits; D044223:Pseudomonas stutzeri / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D004269:DNA, Bacterial; D006841:Hydrocarbons, Aromatic; D009097:Multienzyme Complexes; D021122:Protein Subunits; D011994:Recombinant Proteins; D006899:Mixed Function Oxygenases; D010105:Oxygenases; C114360:toluene 2-xylene monooxygenase; C022336:phenol 2-monooxygenase