PDBsum entry 1c7h

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protein links
Isomerase PDB id
Protein chain
123 a.a. *
Waters ×26
* Residue conservation analysis
PDB id:
Name: Isomerase
Title: Crystal structure of a mutant r75a in ketosteroid isomerase from psedomonas putida biotype b
Structure: Delta-5-3-ketosteroid isomerase. Chain: a. Synonym: ksi, steroid delta-isomerase. Engineered: yes. Mutation: yes
Source: Pseudomonas putida. Organism_taxid: 303. Strain: biotype b. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_cell_line: bl21(de3).
Biol. unit: Dimer (from PDB file)
2.50Å     R-factor:   0.172     R-free:   0.271
Authors: G.H.Nam,D.H.Kim,D.S.Jang,G.Choi,N.C.Ha,B.H.Oh,K.Y.Choi
Key ref:
D.H.Kim et al. (1999). Roles of active site aromatic residues in catalysis by ketosteroid isomerase from Pseudomonas putida biotype B. Biochemistry, 38, 13810-13819. PubMed id: 10529226 DOI: 10.1021/bi991040m
19-Feb-00     Release date:   24-Apr-00    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P07445  (SDIS_PSEPU) -  Steroid Delta-isomerase
131 a.a.
123 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.  - Steroid Delta-isomerase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: A 3-oxo-Delta5-steroid = a 3-oxo-Delta4-steroid
= 3-oxo-Delta(4)-steroid
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     intracellular   1 term 
  Biological process     transport   3 terms 
  Biochemical function     isomerase activity     2 terms  


    Added reference    
DOI no: 10.1021/bi991040m Biochemistry 38:13810-13819 (1999)
PubMed id: 10529226  
Roles of active site aromatic residues in catalysis by ketosteroid isomerase from Pseudomonas putida biotype B.
D.H.Kim, G.H.Nam, D.S.Jang, G.Choi, S.Joo, J.S.Kim, B.H.Oh, K.Y.Choi.
The aromatic residues Phe-54, Phe-82, and Trp-116 in the hydrophobic substrate-binding pocket of Delta(5)-3-ketosteroid isomerase from Pseudomonas putida biotype B have been characterized in their roles in steroid binding and catalysis. Kinetic and equilibrium binding analyses were carried out for the mutant enzymes with the substitutions Phe-54 --> Ala or Leu, Phe-82 --> Ala or Leu, and Trp-116 --> Ala, Phe, or Tyr. The removal of their bulky, aromatic side chains at any of these three positions results in reduced k(cat), particularly when Phe-82 or Trp-116 is replaced by Ala. The results are consistent with the binding interactions of the aromatic residues with the bound steroid contributing to catalysis. All the mutations except the F82A mutation increase K(m); the F82A mutation decreases K(m) by ca. 3-fold, suggesting a possibility that the phenyl ring at position 82 might be unfavorable for substrate binding. The K(D) values for d-equilenin, an intermediate analogue, suggest that a space-filling hydrophobic side chain at position 54, a phenyl ring at position 82, and a nonpolar aromatic or small side chain at position 116 might be favorable for binding the reaction intermediate. In contrast to the increased K(D) for equilenin, the enzymes with any substitutions at positions 54 and 116 display a decreased K(D) for 19-nortestosterone, a product analogue, indicating that Phe-54 and Trp-116 might be unfavorable for product binding. The crystal structure of F82A determined to 2.1-A resolution reveals that Phe-82 is important for maintaining the active site geometry. Taken together, our results demonstrate that Phe-54, Phe-82, and Trp-116 contribute differentially to the stabilization of steroid species including substrate, intermediate, and product.

Literature references that cite this PDB file's key reference

  PubMed id Reference
20143849 W.Childs, and S.G.Boxer (2010).
Proton affinity of the oxyanion hole in the active site of ketosteroid isomerase.
  Biochemistry, 49, 2725-2731.  
19706511 J.P.Schwans, D.A.Kraut, and D.Herschlag (2009).
Determining the catalytic role of remote substrate binding interactions in ketosteroid isomerase.
  Proc Natl Acad Sci U S A, 106, 14271-14275.  
15819891 Y.S.Yun, G.H.Nam, Y.G.Kim, B.H.Oh, and K.Y.Choi (2005).
Small exterior hydrophobic cluster contributes to conformational stability and steroid binding in ketosteroid isomerase from Pseudomonas putida biotype B.
  FEBS J, 272, 1999-2011.
PDB code: 1w6y
12734184 Y.S.Yun, T.H.Lee, G.H.Nam, D.S.Jang, S.Shin, B.H.Oh, and K.Y.Choi (2003).
Origin of the different pH activity profile in two homologous ketosteroid isomerases.
  J Biol Chem, 278, 28229-28236.
PDB code: 1ocv
11274465 D.H.Kim, G.H.Nam, D.S.Jang, S.Yun, G.Choi, H.C.Lee, and K.Y.Choi (2001).
Roles of dimerization in folding and stability of ketosteroid isomerase from Pseudomonas putida biotype B.
  Protein Sci, 10, 741-752.  
10769113 D.H.Kim, D.S.Jang, G.H.Nam, G.Choi, J.S.Kim, N.C.Ha, M.S.Kim, B.H.Oh, and K.Y.Choi (2000).
Contribution of the hydrogen-bond network involving a tyrosine triad in the active site to the structure and function of a highly proficient ketosteroid isomerase from Pseudomonas putida biotype B.
  Biochemistry, 39, 4581-4589.
PDB codes: 1dmm 1dmn 1dmq
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