title: Sequencing and expression of the L-phenylalanine oxidase gene from Pseudomonas sp. P-501. Proteolytic activation of the proenzyme.
abstract: The nucleotide sequence encoding L-phenylalanine oxidase (deaminating and decarboxylating) (PAO) from Pseudomonas sp. P-501 was determined. The open reading frame is arranged in the order of prosequence, alpha subunit, dipeptide and beta subunit from the 5'- to 3'-end. Expression of the gene in Escherichia coli showed that without the prosequence, PAO is produced in small quantity as a soluble form with no visible absorption, but with the prosequence (proPAO), PAO is highly expressed and yellow. The purified proPAO contained one mol of FAD per mol of proPAO polypeptide, but had no catalytic activity. Treatment of proPAO with a mixture of Pronase and trypsin converted the noncatalytic proPAO to the catalytic form, and the Pronase-trypsin-treated proPAO showed kinetic and spectral properties comparable to the native enzyme. These results suggest that in Pseudomonas, PAO is expressed as a proenzyme that is processed by proteolysis to the active form.
authors.affiliation: Division of Biosciences, Graduate School of Fundamental Life Science, School of Science, Kitasato University, Kitasato 1-15-1, Sagamihara, Kanagawa 228-8555, Japan. suzuki@sci.kitasato-u.ac.jp
authors.forename: Haruo
authors.initials: H
authors.lastname: Suzuki
authors.forename: Yuhki
authors.initials: Y
authors.lastname: Higashi
authors.forename: Mai
authors.initials: M
authors.lastname: Asano
authors.forename: Masaya
authors.initials: M
authors.lastname: Suguro
authors.forename: Masayuki
authors.initials: M
authors.lastname: Kigawa
authors.forename: Masahiro
authors.initials: M
authors.lastname: Maeda
authors.forename: Satoshi
authors.initials: S
authors.lastname: Katayama
authors.forename: Etsuko B
authors.initials: EB
authors.lastname: Mukouyama
authors.forename: Koji
authors.initials: K
authors.lastname: Uchiyama
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
chemical_list: D004792:Enzyme Precursors; D000594:Amino Acid Oxidoreductases; C022342:phenylalanine oxidase