PDBsum entry 1ykj

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Oxidoreductase PDB id
Protein chains
394 a.a. *
SO4 ×12
FAD ×2
PHB ×2
PSL ×2
Waters ×542
* Residue conservation analysis
PDB id:
Name: Oxidoreductase
Title: A45g p-hydroxybenzoate hydroxylase with p-hydroxybenzoate bo
Structure: P-hydroxybenzoate hydroxylase. Chain: a, b. Synonym: 4-hydroxybenzoate 3-monooxygenase, phbh. Engineered: yes. Mutation: yes
Source: Pseudomonas aeruginosa. Organism_taxid: 287. Gene: poba. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Dimer (from PQS)
2.00Å     R-factor:   0.201     R-free:   0.265
Authors: L.J.Cole,D.L.Gatti,B.Entsch,D.P.Ballou
Key ref:
L.J.Cole et al. (2005). Removal of a methyl group causes global changes in p-hydroxybenzoate hydroxylase. Biochemistry, 44, 8047-8058. PubMed id: 15924424 DOI: 10.1021/bi050108x
18-Jan-05     Release date:   26-Jul-05    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P20586  (PHHY_PSEAE) -  p-hydroxybenzoate hydroxylase
394 a.a.
394 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 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 = FAD) 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: 10.1021/bi050108x Biochemistry 44:8047-8058 (2005)
PubMed id: 15924424  
Removal of a methyl group causes global changes in p-hydroxybenzoate hydroxylase.
L.J.Cole, D.L.Gatti, B.Entsch, D.P.Ballou.
p-Hydroxybenzoate hydroxylase (PHBH) is a homodimeric flavoprotein monooxygenase that catalyzes the hydroxylation of p-hydroxybenzoate to form 3,4-dihydroxybenzoate. Controlled catalysis is achieved by movement of the flavin and protein between three conformations, in, out, and open [Entsch, B., et al. (2005) Arch. Biochem. Biophys. 433, 297-311]. The open conformation is important for substrate binding and product release, the in conformation for reaction with oxygen and hydroxylation, and the out conformation for the reduction of FAD by NADPH. The open conformation is similar to the structure of Arg220Gln-PHBH in which the backbone peptide loop of residues 43-46, located on the si side of the flavin, is rotated. In this paper, we examine the structure and properties of the Ala45Gly-PHBH mutant enzyme. The crystal structure of the Ala45Gly enzyme is an asymmetric dimer, with one monomer similar (but not identical) to wild-type PHBH, while the other monomer has His72 flipped into solvent and replaced with Glu73 as one of several changes in the structure. The two structures correlate with evidence from kinetic studies for two forms of Ala45Gly-PHBH. One form of the enzyme dominates turnover and hydroxylates, while the other contributes little to turnover and fails to hydroxylate. Ala45Gly-PHBH favors the in conformation over alternative conformations. The effect of this mutation on the structure and function of PHBH illustrates the importance of the si side loop in the conformational state of PHBH and, consequently, the function of the enzyme. This work demonstrates some general principles of how enzymes use conformational movements to allow both access and egress of substrates and product, while restricting access to the solvent at a critical stage in catalysis.