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PDBsum entry 2phh

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protein ligands links
Oxidoreductase PDB id
2phh
Jmol
Contents
Protein chain
391 a.a. *
Ligands
APR
PHB
Waters ×57
* Residue conservation analysis
PDB id:
2phh
Name: Oxidoreductase
Title: The coenzyme analogue adenosine 5-diphosphoribose displaces active site of p-hydroxybenzoate hydroxylase. An x-ray crystallographic investigation
Structure: P-hydroxybenzoate hydroxylase. Chain: a. Engineered: yes
Source: Pseudomonas fluorescens. Organism_taxid: 294
Biol. unit: Dimer (from PQS)
Resolution:
2.70Å     R-factor:   0.168    
Authors: J.M.Van Derlaan,J.Drenth,W.G.J.Hol
Key ref:
J.M.van der Laan et al. (1989). The coenzyme analogue adenosine 5-diphosphoribose displaces FAD in the active site of p-hydroxybenzoate hydroxylase. An x-ray crystallographic investigation. Biochemistry, 28, 7199-7205. PubMed id: 2819062 DOI: 10.1021/bi00444a011
Date:
19-Jun-89     Release date:   15-Oct-90    
PROCHECK
Go to PROCHECK summary
 Headers
 References

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

 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     metabolic process   5 terms 
  Biochemical function     oxidoreductase activity     4 terms  

 

 
DOI no: 10.1021/bi00444a011 Biochemistry 28:7199-7205 (1989)
PubMed id: 2819062  
 
 
The coenzyme analogue adenosine 5-diphosphoribose displaces FAD in the active site of p-hydroxybenzoate hydroxylase. An x-ray crystallographic investigation.
J.M.van der Laan, H.A.Schreuder, M.B.Swarte, R.K.Wierenga, K.H.Kalk, W.G.Hol, J.Drenth.
 
  ABSTRACT  
 
p-Hydroxybenzoate hydroxylase (PHBH) is an NADPH-dependent enzyme. To locate the NADPH binding site, the enzyme was crystallized under anaerobic conditions in the presence of the substrate p-hydroxybenzoate, the coenzyme analogue adenosine 5-diphosphoribose (ADPR), and sodium dithionite. This yielded colorless crystals that were suitable for X-ray analysis. Diffraction data were collected up to 2.7-A resolution. A difference Fourier between data from these colorless crystals and data from yellow crystals of the enzyme-substrate complex showed that in the colorless crystals the flavin ring was absent. The adenosine 5'-diphosphate moiety, which is the common part between FAD and ADPR, was still present. After restrained least-squares refinement of the enzyme-substrate complex with the riboflavin omitted from the model, additional electron density appeared near the pyrophosphate, which indicated the presence of an ADPR molecule in the FAD binding site of PHBH. The complete ADPR molecule was fitted to the electron density, and subsequent least-squares refinement resulted in a final R factor of 16.8%. Replacement of bound FAD by ADPR was confirmed by equilibrium dialysis, where it was shown that ADPR can effectively remove FAD from the enzyme under mild conditions in 0.1 M potassium phosphate buffer, pH 8.0. The empty pocket left by the flavin ring is filled by solvent, leaving the architecture of the active site and the binding of the substrate largely unaffected.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
9694855 M.H.Eppink, H.A.Schreuder, and W.J.van Berkel (1998).
Interdomain binding of NADPH in p-hydroxybenzoate hydroxylase as suggested by kinetic, crystallographic and modeling studies of histidine 162 and arginine 269 variants.
  J Biol Chem, 273, 21031-21039.
PDB codes: 1bgj 1bgn
  9385648 M.H.Eppink, H.A.Schreuder, and W.J.Van Berkel (1997).
Identification of a novel conserved sequence motif in flavoprotein hydroxylases with a putative dual function in FAD/NAD(P)H binding.
  Protein Sci, 6, 2454-2458.  
8706756 B.Seibold, M.Matthes, M.H.Eppink, F.Lingens, W.J.Van Berkel, and R.Müller (1996).
4-Hydroxybenzoate hydroxylase from Pseudomonas sp. CBS3. Purification, characterization, gene cloning, sequence analysis and assignment of structural features determining the coenzyme specificity.
  Eur J Biochem, 239, 469-478.  
7628466 M.H.Eppink, H.A.Schreuder, and W.J.Van Berkel (1995).
Structure and function of mutant Arg44Lys of 4-hydroxybenzoate hydroxylase implications for NADPH binding.
  Eur J Biochem, 231, 157-165.
PDB code: 1bkw
  1729209 B.Averhoff, L.Gregg-Jolly, D.Elsemore, and L.N.Ornston (1992).
Genetic analysis of supraoperonic clustering by use of natural transformation in Acinetobacter calcoaceticus.
  J Bacteriol, 174, 200-204.  
1409567 H.A.Schreuder, J.M.van der Laan, M.B.Swarte, K.H.Kalk, W.G.Hol, and J.Drenth (1992).
Crystal structure of the reduced form of p-hydroxybenzoate hydroxylase refined at 2.3 A resolution.
  Proteins, 14, 178-190.  
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.