UniProt functional annotation for P00438



	UniProt functional annotation for P00438

UniProt code: P00438.

Organism: Pseudomonas fluorescens.

Taxonomy: Bacteria; Proteobacteria; Gammaproteobacteria; Pseudomonadales; Pseudomonadaceae; Pseudomonas.

Function: Catalyzes the incorporation of an atom of dioxygen into p- hydroxybenzoate (p-OHB) to form 3,4-dihydroxybenzoate (3,4DOHB). The reaction occurs in two parts: reduction of the flavin adenine dinucleotide (FAD) in the enzyme by reduced nicotinamide adenine dinucleotide phosphate (NADPH) in response to binding p-hydroxybenzoate to the enzyme and oxidation of reduced FAD with oxygen to form a hydroperoxide, which then oxygenates p-hydroxybenzoate. {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:1459126, ECO:0000269|PubMed:2819062, ECO:0000269|PubMed:3351945, ECO:0000269|PubMed:7520279, ECO:0000269|PubMed:7628466, ECO:0000269|PubMed:7756982, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855}.

Catalytic activity: Reaction=4-hydroxybenzoate + H(+) + NADPH + O2 = 3,4-dihydroxybenzoate + H2O + NADP(+); Xref=Rhea:RHEA:19477, ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:15379, ChEBI:CHEBI:17879, ChEBI:CHEBI:36241, ChEBI:CHEBI:57783, ChEBI:CHEBI:58349; EC=1.14.13.2; Evidence={ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:1459126, ECO:0000269|PubMed:7628466, ECO:0000269|PubMed:7756982, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855};

Cofactor: Name=FAD; Xref=ChEBI:CHEBI:57692; Evidence={ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:1409567, ECO:0000269|PubMed:1459126, ECO:0000269|PubMed:2819062, ECO:0000269|PubMed:3351945, ECO:0000269|PubMed:7628466, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855}; Note=Binds 1 FAD per subunit. {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:2819062, ECO:0000269|PubMed:3351945, ECO:0000269|PubMed:7628466, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855};

Biophysicochemical properties: Kinetic parameters: KM=15 uM for p-OHB {ECO:0000269|PubMed:9578477}; KM=15 uM for p-OHB (at pH 6) {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859}; KM=20 uM for p-OHB {ECO:0000269|PubMed:7756982, ECO:0000269|PubMed:9694855}; KM=25 uM for p-OHB (at 25 degrees Celsius) {ECO:0000269|PubMed:1459126}; KM=30 uM for NADPH {ECO:0000269|PubMed:9694855}; KM=30 uM for p-OHB (at 6 degrees Celsius) {ECO:0000269|PubMed:1459126}; KM=34 uM for NADPH (at pH 6) {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859}; KM=40 uM for NADPH (at 6 degrees Celsius) {ECO:0000269|PubMed:1459126}; KM=50 uM for NADPH (at 25 degrees Celsius) {ECO:0000269|PubMed:1459126}; KM=50 uM for NADPH {ECO:0000269|PubMed:9578477}; KM=70 uM for NADPH {ECO:0000269|PubMed:7756982}; Note=Kcat is 55 sec(-1) for hydroxylase activity (PubMed:9578477, PubMed:9694855). Kcat is 55 sec(-1) for hydroxylase activity (at 25 degrees Celsius) (PubMed:1459126). Kcat is 55 sec(-1) for hydroxylase activity (at pH 8) (PubMed:10025942, PubMed:10493859). Kcat is 9 sec(-1) for hydroxylase activity (at 6 degrees Celsius) (PubMed:1459126). {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:1459126, ECO:0000269|PubMed:7756982, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855};

Pathway: Aromatic compound metabolism; benzoate degradation via hydroxylation; 3,4-dihydroxybenzoate from benzoate: step 2/2. {ECO:0000305}.

Subunit: Homodimer. {ECO:0000269|PubMed:10025942, ECO:0000269|PubMed:10493859, ECO:0000269|PubMed:2553983, ECO:0000269|PubMed:2819062, ECO:0000269|PubMed:3351945, ECO:0000269|PubMed:40036, ECO:0000269|PubMed:7520279, ECO:0000269|PubMed:7628466, ECO:0000269|PubMed:7756982, ECO:0000269|PubMed:9578477, ECO:0000269|PubMed:9694855}.

Similarity: Belongs to the aromatic-ring hydroxylase family. {ECO:0000305}.

Annotations taken from UniProtKB at the EBI.


Organism:		Pseudomonas fluorescens.
Taxonomy:		Bacteria; Proteobacteria; Gammaproteobacteria; Pseudomonadales; Pseudomonadaceae; Pseudomonas.



Function:		Catalyzes the incorporation of an atom of dioxygen into p- hydroxybenzoate (p-OHB) to form 3,4-dihydroxybenzoate (3,4DOHB). The reaction occurs in two parts: reduction of the flavin adenine dinucleotide (FAD) in the enzyme by reduced nicotinamide adenine dinucleotide phosphate (NADPH) in response to binding p-hydroxybenzoate to the enzyme and oxidation of reduced FAD with oxygen to form a hydroperoxide, which then oxygenates p-hydroxybenzoate. {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:1459126, ECO:0000269\|PubMed:2819062, ECO:0000269\|PubMed:3351945, ECO:0000269\|PubMed:7520279, ECO:0000269\|PubMed:7628466, ECO:0000269\|PubMed:7756982, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855}.


Catalytic activity:		Reaction=4-hydroxybenzoate + H(+) + NADPH + O2 = 3,4-dihydroxybenzoate + H2O + NADP(+); Xref=Rhea:RHEA:19477, ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:15379, ChEBI:CHEBI:17879, ChEBI:CHEBI:36241, ChEBI:CHEBI:57783, ChEBI:CHEBI:58349; EC=1.14.13.2; Evidence={ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:1459126, ECO:0000269\|PubMed:7628466, ECO:0000269\|PubMed:7756982, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855};
Cofactor:		Name=FAD; Xref=ChEBI:CHEBI:57692; Evidence={ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:1409567, ECO:0000269\|PubMed:1459126, ECO:0000269\|PubMed:2819062, ECO:0000269\|PubMed:3351945, ECO:0000269\|PubMed:7628466, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855}; Note=Binds 1 FAD per subunit. {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:2819062, ECO:0000269\|PubMed:3351945, ECO:0000269\|PubMed:7628466, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855};
Biophysicochemical properties:		Kinetic parameters: KM=15 uM for p-OHB {ECO:0000269\|PubMed:9578477}; KM=15 uM for p-OHB (at pH 6) {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859}; KM=20 uM for p-OHB {ECO:0000269\|PubMed:7756982, ECO:0000269\|PubMed:9694855}; KM=25 uM for p-OHB (at 25 degrees Celsius) {ECO:0000269\|PubMed:1459126}; KM=30 uM for NADPH {ECO:0000269\|PubMed:9694855}; KM=30 uM for p-OHB (at 6 degrees Celsius) {ECO:0000269\|PubMed:1459126}; KM=34 uM for NADPH (at pH 6) {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859}; KM=40 uM for NADPH (at 6 degrees Celsius) {ECO:0000269\|PubMed:1459126}; KM=50 uM for NADPH (at 25 degrees Celsius) {ECO:0000269\|PubMed:1459126}; KM=50 uM for NADPH {ECO:0000269\|PubMed:9578477}; KM=70 uM for NADPH {ECO:0000269\|PubMed:7756982}; Note=Kcat is 55 sec(-1) for hydroxylase activity (PubMed:9578477, PubMed:9694855). Kcat is 55 sec(-1) for hydroxylase activity (at 25 degrees Celsius) (PubMed:1459126). Kcat is 55 sec(-1) for hydroxylase activity (at pH 8) (PubMed:10025942, PubMed:10493859). Kcat is 9 sec(-1) for hydroxylase activity (at 6 degrees Celsius) (PubMed:1459126). {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:1459126, ECO:0000269\|PubMed:7756982, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855};
Pathway:		Aromatic compound metabolism; benzoate degradation via hydroxylation; 3,4-dihydroxybenzoate from benzoate: step 2/2. {ECO:0000305}.
Subunit:		Homodimer. {ECO:0000269\|PubMed:10025942, ECO:0000269\|PubMed:10493859, ECO:0000269\|PubMed:2553983, ECO:0000269\|PubMed:2819062, ECO:0000269\|PubMed:3351945, ECO:0000269\|PubMed:40036, ECO:0000269\|PubMed:7520279, ECO:0000269\|PubMed:7628466, ECO:0000269\|PubMed:7756982, ECO:0000269\|PubMed:9578477, ECO:0000269\|PubMed:9694855}.
Similarity:		Belongs to the aromatic-ring hydroxylase family. {ECO:0000305}.