According to the literature [10], [13], [14] & [40], this enzyme catalyzes the following reactions: (A) Hydride transfer from NADPH to FAD, forming FADH2 (Reduction of FAD by NADPH) (Reductive half-reaction): (B) Oxygenation of hydroxybenzoate by O2 at FADH2 (Oxidative half-reaction): (B1) Oxygenation to form a flavin 4a-hydroperoxide intermediate. (B2) Additive double-bond deformation; Addition of hydroxyl group to substrate, 4-hydroxybenzoate, by flavin 4a-hydroperoxide intermediate (Hydroxylation of substrate), forming 3,4-dihydroxybenzoate and a flavin 4a-hydroxide intermediate. (B3) Eliminative double-bond formation; Elimination of H2O from flavin 4a-hydroxide intermediate, giving FAD.
Crystallization and preliminary x-ray investigation of p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens.
Analysis of the active site of the flavoprotein p-hydroxybenzoate hydroxylase and some ideas with respect to its reaction mechanism.
Crystal structure of the reduced form of p-hydroxybenzoate hydroxylase refined at 2.3 A resolution.
X-RAY CRYSTALLOGRAPHY (2.3 ANGSTROMS) OF REDUCED FORM
Role of Tyr201 and Tyr385 in substrate activation by p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens.
Crystal structures of wild-type p-hydroxybenzoate hydroxylase complexed with 4-aminobenzoate,2,4-dihydroxybenzoate, and 2-hydroxy-4-aminobenzoate and of the Tyr222Ala mutant complexed with 2-hydroxy-4-aminobenzoate. Evidence for a proton channel and a new binding mode of the flavin ring.
Crystal structure of p-hydroxybenzoate hydroxylase reconstituted with the modified FAD present in alcohol oxidase from methylotrophic yeasts: evidence for an arabinoflavin.
The mobile flavin of 4-OH benzoate hydroxylase.
Structure and function of mutant Arg44Lys of 4-hydroxybenzoate hydroxylase implications for NADPH binding.
X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF MUTANT LYS-44
pH-dependent structural changes in the active site of p-hydroxybenzoate hydroxylase point to the importance of proton and water movements during catalysis.
19F NMR study on the regiospecificity of hydroxylation of tetrafluoro-4-hydroxybenzoate by wild-type and Y385F p-hydroxybenzoate hydroxylase: evidence for a consecutive oxygenolytic dehalogenation mechanism.
Electron microscopic studies on microcrystals of parahydroxybenzoate hydroxylase from Pseudomonas fluorescens.
Identification of a novel conserved sequence motif in flavoprotein hydroxylases with a putative dual function in FAD/NAD(P)H binding.
Lys42 and Ser42 variants of p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens reveal that Arg42 is essential for NADPH binding.
X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF MUTANT LYS-42 AND SER-42
Interdomain binding of NADPH in p-hydroxybenzoate hydroxylase as suggested by kinetic, crystallographic and modeling studies of histidine 162 and arginine 269 variants.
Mechanistic insights into p-hydroxybenzoate hydroxylase from studies of the mutant Ser212Ala.
Structure-function correlations of the reaction of reduced nicotinamide analogues with p-hydroxybenzoate hydroxylase substituted with a series of 8-substituted flavins.
Substrate recognition by "password" in p-hydroxybenzoate hydroxylase.
Using Raman spectroscopy to monitor the solvent-exposed and "buried" forms of flavin in p-hydroxybenzoate hydroxylase.
Phe161 and Arg166 variants of p-hydroxybenzoate hydroxylase. Implications for NADPH recognition and structural stability.
X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF VARIANTS
Switch of coenzyme specificity of p-hydroxybenzoate hydroxylase.
Crystal structure of p-hydroxybenzoate hydroxylase.
Combined quantum mechanical and molecular mechanical reaction pathway calculation for aromatic hydroxylation by p-hydroxybenzoate-3-hydroxylase.
Modelling flavin and substrate substituent effects on the activation barrier and rate of oxygen transfer by p-hydroxybenzoate hydroxylase.
Protein dynamics control proton transfers to the substrate on the His72Asn mutant of p-hydroxybenzoate hydroxylase.
A rate-limiting conformational change of the flavin in p-hydroxybenzoate hydroxylase is necessary for ligand exchange and catalysis: studies with 8-mercapto- and 8-hydroxy-flavins.
Comparing protein-ligand interactions in solution and single crystals by Raman spectroscopy.
Protein and ligand dynamics in 4-hydroxybenzoate hydroxylase.
Conformational changes combined with charge-transfer interactions are essential for reduction in catalysis by p-hydroxybenzoate hydroxylase.
Altered balance of half-reactions in p-hydroxybenzoate hydroxylase caused by substituting the 2'-carbon of FAD with fluorine.
Increased positive electrostatic potential in p-hydroxybenzoate hydroxylase accelerates hydroxylation but slows turnover.
Model studies on p-hydroxybenzoate hydroxylase. The catalytic role of Arg-214 and Tyr-201 in the hydroxylation step.
Reaction mechanism of flavin-dependent hydroxylation. Evolution of a non-imitable enzyme.
Protein dynamics and electrostatics in the function of p-hydroxybenzoate hydroxylase.
Evolution of biocatalysis 4. Nicotinamide, flavin and dioxygen dependent hydroxylation. Origin of a non-imitable enzyme.
Molecular modeling reveals the possible importance of a carbonyl oxygen binding pocket for the catalytic mechanism of p-hydroxybenzoate hydroxylase.
Crystal structure of p-hydroxybenzoate hydroxylase complexed with its reaction product 3,4-dihydroxybenzoate.
The coenzyme analogue adenosine 5-diphosphoribose displaces FAD in the active site of p-hydroxybenzoate hydroxylase. An x-ray crystallographic investigation.
Crystal structure of the p-hydroxybenzoate hydroxylase-substrate complex refined at 1.9 A resolution. Analysis of the enzyme-substrate and enzyme-product complexes.
Crystal structures of mutant Pseudomonas aeruginosa p-hydroxybenzoate hydroxylases: the Tyr201Phe, Tyr385Phe, and Asn300Asp variants.
X-RAY CRYSTALLOGRAPHY (2.3 ANGSTROMS) OF MUTANTS