PDBsum entry 1pdh

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Oxidoreductase PDB id
Protein chain
391 a.a. *
Waters ×284
* Residue conservation analysis
PDB id:
Name: Oxidoreductase
Title: Crystal structure of p-hydroxybenzoate hydroxylase reconstituted with the modified fad present in alcohol oxidase from methylotrophic yeasts: evidence for an arabinoflavin
Structure: P-hydroxybenzoate hydroxylase. Chain: a. Engineered: yes
Source: Pseudomonas fluorescens. Organism_taxid: 294
Biol. unit: Dimer (from PQS)
2.10Å     R-factor:   0.179    
Authors: H.A.Schreuder,M.H.M.Eppink,W.J.H.Van Berkel
Key ref:
W.J.van Berkel et al. (1994). Crystal structure of p-hydroxybenzoate hydroxylase reconstituted with the modified FAD present in alcohol oxidase from methylotrophic yeasts: evidence for an arabinoflavin. Protein Sci, 3, 2245-2253. PubMed id: 7756982 DOI: 10.1002/pro.5560031210
01-Dec-94     Release date:   31-Mar-95    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P00438  (PHHY_PSEFL) -  p-hydroxybenzoate hydroxylase
394 a.a.
391 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.  - 4-hydroxybenzoate 3-monooxygenase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

Benzoate Metabolism
      Reaction: 4-hydroxybenzoate + NADPH + O2 = protocatechuate + NADP+ + H2O
Bound ligand (Het Group name = PHB)
corresponds exactly
+ O(2)
= protocatechuate
+ NADP(+)
+ H(2)O
      Cofactor: FAD
Bound ligand (Het Group name = FAS) corresponds exactly
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     metabolic process   5 terms 
  Biochemical function     oxidoreductase activity     4 terms  


DOI no: 10.1002/pro.5560031210 Protein Sci 3:2245-2253 (1994)
PubMed id: 7756982  
Crystal structure of p-hydroxybenzoate hydroxylase reconstituted with the modified FAD present in alcohol oxidase from methylotrophic yeasts: evidence for an arabinoflavin.
W.J.van Berkel, M.H.Eppink, H.A.Schreuder.
The flavin prosthetic group (FAD) of p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens was replaced by a stereochemical analog, which is spontaneously formed from natural FAD in alcohol oxidases from methylotrophic yeasts. Reconstitution of p-hydroxybenzoate hydroxylase from apoprotein and modified FAD is a rapid process complete within seconds. Crystals of the enzyme-substrate complex of modified FAD-containing p-hydroxybenzoate hydroxylase diffract to 2.1 A resolution. The crystal structure provides direct evidence for the presence of an arabityl sugar chain in the modified form of FAD. The isoalloxazine ring of the arabinoflavin adenine dinucleotide (a-FAD) is located in a cleft outside the active site as recently observed in several other p-hydroxybenzoate hydroxylase complexes. Like the native enzyme, a-FAD-containing p-hydroxybenzoate hydroxylase preferentially binds the phenolate form of the substrate (pKo = 7.2). The substrate acts as an effector highly stimulating the rate of enzyme reduction by NADPH (kred > 500 s-1). The oxidative part of the catalytic cycle of a-FAD-containing p-hydroxybenzoate hydroxylase differs from native enzyme. Partial uncoupling of hydroxylation results in the formation of about 0.3 mol of 3,4-dihydroxybenzoate and 0.7 mol of hydrogen peroxide per mol NADPH oxidized. It is proposed that flavin motion in p-hydroxybenzoate hydroxylase is important for efficient reduction and that the flavin "out" conformation is associated with the oxidase activity.
  Selected figure(s)  
Figure 3.
Fig. 3. Superposition of thestructurs of hea-FAD-containingenzyme-4hydroxy- benzoatecomplexandthenative enzyme- 2,4-dihydroxybenzoatecomplex.The structure of the4-hydroxybenzoatecom- plexwitha-FADisdrawnwithsolid bonds;thestructure of the 2,4-dihydroxy- benzoatecomplexwithnaturalFAD s drawnwithopenbonds.The view is from theflavinringtowardtheribitylchain. Brokenlinesindicatethehydrogenbond between the02' hydroxyl groupandthe OEl of Gln 102, is presentinboth complexes.
Figure 5.
Fig. 5. Stereodiagram of hesuperosi- ion of native2,4-dihydroxybenzoate complexedp-hydroxybenzoate hydroxylase Schreuderetal., 1994; graybonds)and hea-FAD-containingenzyme-substrate complex(blackbonds).Theflavinring ccupiesthe``out''postioninbothcom- lexes, buttheflavinring is slightly fur- heroutinthe2,4-dihydroxybenzoate complex,presumably because of ahydro- genbondcontact,indicatedbyabroken line,between the 2-hydroxygroup of he substrateanalogandthe N3 of thelavin ring.
  The above figures are reprinted from an Open Access publication published by the Protein Society: Protein Sci (1994, 3, 2245-2253) copyright 1994.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21402075 G.Volkers, G.J.Palm, M.S.Weiss, G.D.Wright, and W.Hinrichs (2011).
Structural basis for a new tetracycline resistance mechanism relying on the TetX monooxygenase.
  FEBS Lett, 585, 1061-1066.
PDB codes: 2xdo 2xyo 2y6q 2y6r
19348008 G.Tabacchi, D.Zucchini, G.Caprini, A.Gamba, F.Lederer, M.A.Vanoni, and E.Fois (2009).
L-lactate dehydrogenation in flavocytochrome b2: a first principles molecular dynamics study.
  FEBS J, 276, 2368-2380.  
18176933 E.V.Kudryashova, A.J.Visser, and W.J.van Berkel (2008).
Monomer formation and function of p-hydroxybenzoate hydroxylase in reverse micelles and in dimethylsulfoxide/water mixtures.
  Chembiochem, 9, 413-419.  
14622288 M.H.Hefti, J.Vervoort, and W.J.van Berkel (2003).
Deflavination and reconstitution of flavoproteins.
  Eur J Biochem, 270, 4227-4242.  
12081493 B.A.Palfey, R.Basu, K.K.Frederick, B.Entsch, and D.P.Ballou (2002).
Role of protein flexibility in the catalytic cycle of p-hydroxybenzoate hydroxylase elucidated by the Pro293Ser mutant.
  Biochemistry, 41, 8438-8446.  
11805318 J.Wang, M.Ortiz-Maldonado, B.Entsch, V.Massey, D.Ballou, and D.L.Gatti (2002).
Protein and ligand dynamics in 4-hydroxybenzoate hydroxylase.
  Proc Natl Acad Sci U S A, 99, 608-613.
PDB codes: 1k0i 1k0j 1k0l
11248022 M.D.Altose, Y.Zheng, J.Dong, B.A.Palfey, and P.R.Carey (2001).
Comparing protein-ligand interactions in solution and single crystals by Raman spectroscopy.
  Proc Natl Acad Sci U S A, 98, 3006-3011.  
11082194 M.H.Eppink, E.Cammaart, D.Van Wassenaar, W.J.Middelhoven, and W.J.van Berkel (2000).
Purification and properties of hydroquinone hydroxylase, a FAD-dependent monooxygenase involved in the catabolism of 4-hydroxybenzoate in Candida parapsilosis CBS604.
  Eur J Biochem, 267, 6832-6840.  
10600126 M.Ortiz-Maldonado, D.Gatti, D.P.Ballou, and V.Massey (1999).
Structure-function correlations of the reaction of reduced nicotinamide analogues with p-hydroxybenzoate hydroxylase substituted with a series of 8-substituted flavins.
  Biochemistry, 38, 16636-16647.
PDB code: 1d7l
10606503 Y.Zheng, J.Dong, B.A.Palfey, and P.R.Carey (1999).
Using Raman spectroscopy to monitor the solvent-exposed and "buried" forms of flavin in p-hydroxybenzoate hydroxylase.
  Biochemistry, 38, 16727-16732.  
9200706 G.R.Moran, B.Entsch, B.A.Palfey, and D.P.Ballou (1997).
Electrostatic effects on substrate activation in para-hydroxybenzoate hydroxylase: studies of the mutant lysine 297 methionine.
  Biochemistry, 36, 7548-7556.  
8647102 F.J.van der Bolt, J.Vervoort, and W.J.van Berkel (1996).
Flavin motion in p-hydroxybenzoate hydroxylase. Substrate and effector specificity of the Tyr22-->Ala mutant.
  Eur J Biochem, 237, 592-600.  
8703933 G.R.Moran, B.Entsch, B.A.Palfey, and D.P.Ballou (1996).
Evidence for flavin movement in the function of p-hydroxybenzoate hydroxylase from studies of the mutant Arg220Lys.
  Biochemistry, 35, 9278-9285.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.