Cloned (Comment) | Organism |
---|---|
gene phhA, sequence comparisons and phylogenetic analysis and tree | Aspergillus niger |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
4-hydroxybenzoate + NADPH + H+ + O2 | Aspergillus niger | - |
3,4-dihydroxybenzoate + NADP+ + H2O | - |
? | |
4-hydroxybenzoate + NADPH + H+ + O2 | Aspergillus niger N402 | - |
3,4-dihydroxybenzoate + NADP+ + H2O | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Aspergillus niger | - |
- |
- |
Aspergillus niger N402 | - |
- |
- |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
4-hydroxybenzoate + NADPH + H+ + O2 = 3,4-dihydroxybenzoate + NADP+ + H2O | the catalytic mechanism of PHBH from Pseudomonas species includes the reductive half-reaction, in which 4-hydroxybenzoate (4-HB) and some substrate analogues can act as effectors by stimulating the two-electron reduction of the FAD cofactor by NAD(P)H. In the oxidative half-reaction, the enzyme activates molecular oxygen through formation of a transiently stable flavin C4a-hydroperoxide and subsequently regioselectively incorporates one oxygen atom into the substrate. Based on the structural and kinetic properties of wild-type enzyme and a range of protein variants, the catalytic cycle of PHBH has been proposed to involve flavin movements in and out of the active site and opening and closure of a substrate transport channel. The C3 atom of the aromatic ring of the substrate is in close vicinity of the C4a atom of the isoalloxazine ring of FAD, which occupies the conformation. This active site configuration allows the attack of the flavin C4a-hydroperoxide oxygenation species onto the C3 atom of the substrate. The electrophilic ortho-hydroxylation reaction is stimulated by deprotonation of the phenolic moiety of 4-hydroxybenzoate, as facilitated by a water involved proton relay network that connects Tyr201 via Tyr385 and His72 with the protein surface | Aspergillus niger |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
4-hydroxybenzoate + NADPH + H+ + O2 | - |
Aspergillus niger | 3,4-dihydroxybenzoate + NADP+ + H2O | - |
? | |
4-hydroxybenzoate + NADPH + H+ + O2 | - |
Aspergillus niger N402 | 3,4-dihydroxybenzoate + NADP+ + H2O | - |
? | |
additional information | substrate binding analysis and structure, each subunit of the dimeric enzyme contains a split substrate-binding domain, overview | Aspergillus niger | ? | - |
- |
|
additional information | substrate binding analysis and structure, each subunit of the dimeric enzyme contains a split substrate-binding domain, overview | Aspergillus niger N402 | ? | - |
- |
Subunits | Comment | Organism |
---|---|---|
homodimer | each subunit of the dimeric enzyme contains a split FAD-binding domain and a split substrate-binding domain | Aspergillus niger |
More | structure modeling of the PhhA subunit, the FAD-binding domain, the FAD cofactor, the substrate-binding domain, the thioredoxin domain, and substrate 4-hydroxybenzoate, overview | Aspergillus niger |
Synonyms | Comment | Organism |
---|---|---|
4-hydroxybenzoate 3-hydroxylase | - |
Aspergillus niger |
An_PhhA | - |
Aspergillus niger |
FAD-dependent 4-hydroxybenzoate hydroxylase | - |
Aspergillus niger |
fungal 4-hydroxybenzoate 3-hydroxylase | - |
Aspergillus niger |
PHBH | - |
Aspergillus niger |
phhA | - |
Aspergillus niger |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Aspergillus niger |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.6 | - |
assay at | Aspergillus niger |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
FAD | each subunit of the dimeric enzyme contains a split FAD-binding domain | Aspergillus niger | |
NADPH | - |
Aspergillus niger |
Organism | Comment | Expression |
---|---|---|
Aspergillus niger | the enzyme is induced by 4-hydroxybenzoate as carbon source | up |
General Information | Comment | Organism |
---|---|---|
evolution | evolutionary relationship between the NAD(P)H-dependent FAD-containing 4-hydroxybenzoate hydroxylases and phylogenetic analysis of group A FPMOs, overview | Aspergillus niger |
evolution | phylogenetic analysis shows that FAD-dependent 4-hydroxybenzoate hydroxylases reside in distinct clades of the group A flavoprotein monooxygenase (FPMO) family, indicating their separate divergence from a common ancestor. Protein homology modeling reveals that the fungal 4-hydroxybenzoate 3-hydroxylase PhhA is structurally related to phenol hydroxylase (PHHY) and 3-hydroxybenzoate 4-hydroxylase (3HB4H). 4-Hydroxybenzoate 1-hydroxylase (4HB1H) from yeast catalyzes an oxidative decarboxylation reaction and is structurally similar to 3-hydroxybenzoate 6-hydroxylase (3HB6H), salicylate hydroxylase (SALH) and 6-hydroxynicotinate 3-monooxygenase (6HNMO). Group A FPMOs are involved in the aerobic microbial catabolism of 4-hydroxybenzoate. Phylogenetic analysis and structure comparisons, detailed overview | Aspergillus niger |
metabolism | enzyme PHBH is almost exclusively found in prokaryotes, where its induction, usually as a consequence of lignin degradation, results in the regioselective formation of protocatechuate, one of the central intermediates in the global carbon cycle | Aspergillus niger |
additional information | enzyme protein homology modeling of An_PhhA using the structure file of Tc_PHHY (PDB ID 1pn0) as template, overview. Hydroxylase enzymes structure comparisons, overview | Aspergillus niger |
physiological function | 4-hydroxybenzoate (4-HB) is a common intermediate in lignin degradation. It is one of the aromatic acids that arise from the Calpha-Cbeta cleavage of lignin components. In aerobic bacteria, 4-HB usually is converted to the ring-fission substrate 3,4-dihydroxybenzoate (protocatechuate, PCA). This reaction is catalyzed by the NAD(P)H-dependent flavoprotein monooxygenase (FPMO) 4-hydroxybenzoate 3-hydroxylase (PHBH) | Aspergillus niger |