Sequence and organization of pobA, the gene coding for p-hydroxybenzoate hydroxylase, an inducible enzyme from Pseudomonas aeruginosa.
The only recognized gene for the metabolism of p-hydroxybenzoate in Pseudomonads (pobA) has been isolated from Pseudomonas aeruginosa to provide the DNA for mutagenesis studies of the protein product, p-hydroxybenzoate hydroxylase. Since pobA is induced by p-hydroxybenzoate to produce large amounts of enzyme, its regulation in P. aeruginosa is significant. The nucleotide sequence of pobA is presented with the derived amino acid (aa) sequence, which has only two substitutions compared to the amino acid sequence obtained from the enzyme from P. fluorescens. The derived amino acid sequence predicts that the enzyme is a single polypeptide of 394 aa residues and contains one molecule of FAD. The complete structure of the protein from P. aeruginosa can be derived by analogy from the published structure of the protein from P. fluorescens. Transcription mapping was used to determine that there is one site for the initiation of mRNA synthesis in P. aeruginosa. The presence of a putative operator in the sequence suggests primary regulation by a repressor protein which binds p-hydroxybenzoate. The ribosome-binding site permits translation of the gene in Escherichia coli at levels comparable to its production in P. aeruginosa, but it produces no detectable product in E. coli under the influence of its own promoter sequence. The promoter does not conform to the common consensus sequence of E. coli promoters. The results have identified an apparent novel promoter for P. aeruginosa, which may reflect the presence of a sigma factor required for pobA induction. Repression of expression by glucose suggests a binding site in the sequence for catabolite repression.
Entsch
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B
Department of Biochemistry, Microbiology and Nutrition, University of New England, Armidale, N.S.W., Australia.
Nan
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Y
Weaich
K
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Scott
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KF
D015109:4-Hydroxybenzoate-3-Monooxygenase / Q000096:biosynthesis / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003470:Culture Media; D004790:Enzyme Induction; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D006899:Mixed Function Oxygenases / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids; D011550:Pseudomonas aeruginosa / Q000235:genetics*; D012313:RNA / Q000302:isolation & purification; D014158:Transcription, Genetic; D014169:Transformation, Bacterial
D003470:Culture Media; D012313:RNA; D006899:Mixed Function Oxygenases; D015109:4-Hydroxybenzoate-3-Monooxygenase