PDBsum entry 1grp

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Oxidoreductase (NAD(a)-choh(d)) PDB id
Protein chain
414 a.a. *
* Residue conservation analysis
PDB id:
Name: Oxidoreductase (NAD(a)-choh(d))
Title: Regulatory and catalytic mechanisms in escherichia coli isoc dehydrogenase: multiple roles for n115
Structure: Isocitrate dehydrogenase. Chain: a. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562
Biol. unit: Dimer (from PQS)
2.50Å     R-factor:   0.195    
Authors: J.A.Grobler,J.H.Hurley
Key ref:
R.Chen et al. (1996). Second-site suppression of regulatory phosphorylation in Escherichia coli isocitrate dehydrogenase. Protein Sci, 5, 287-295. PubMed id: 8745407 DOI: 10.1002/pro.5560050213
12-Oct-95     Release date:   03-Apr-96    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P08200  (IDH_ECOLI) -  Isocitrate dehydrogenase [NADP]
416 a.a.
414 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.  - Isocitrate dehydrogenase (NADP(+)).
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

Citric acid cycle
      Reaction: Isocitrate + NADP+ = 2-oxoglutarate + CO2 + NADPH
Bound ligand (Het Group name = ICT)
corresponds exactly
+ NADP(+)
= 2-oxoglutarate
+ CO(2)
      Cofactor: Mn(2+) or Mg(2+)
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   1 term 
  Biological process     oxidation-reduction process   5 terms 
  Biochemical function     oxidoreductase activity     6 terms  


DOI no: 10.1002/pro.5560050213 Protein Sci 5:287-295 (1996)
PubMed id: 8745407  
Second-site suppression of regulatory phosphorylation in Escherichia coli isocitrate dehydrogenase.
R.Chen, J.A.Grobler, J.H.Hurley, A.M.Dean.
Inactivation of Escherichia coli isocitrate dehydrogenase upon phosphorylation at S113 depends upon the direct electrostatic repulsion of the negatively charged gamma-carboxylate of isocitrate by the negatively charged phosphoserine. The effect is mimicked by replacing S113 with aspartate or glutamate, which reduce performance (kcat/K(i).isocitrat/ Km.NADP) by a factor of 10(7). Here, we demonstrate that the inactivating effects of the electrostatic repulsion are completely eliminated by a second-site mutation, and provide the structural basis for this striking example of intragenic suppression. N115 is adjacent to S113 on one face of the D-helix, interacts with isocitrate and NADP+, and has been postulated to serve in both substrate binding and in catalysis. The single N115L substitution reduces affinity for isocitrate by a factor of 50 and performance by a factor of 500. However, the N115L substitution completely suppresses the inactivating electrostatic effects of S113D or S113E: the performance of the double mutants is 10(5) higher than the S113D and S113E single mutants. These mutations have little effect on the kinetics of alternative substrates, which lack the charged gamma-carboxylate of isocitrate. Both glutamate and aspartate at site 113 remain fully ionized in the presence of leucine. In the crystal structure of the N115L mutant, the leucine adopts a different conformer from the wild-type asparagine. Repacking around the leucine forces the amino-terminus of the D-helix away from the rest of the active site. The hydrogen bond between E113 and N115 in the S113E single mutant is broken in the S113E/N115L mutant, allowing the glutamate side chain to move away from the gamma-carboxylate of isocitrate. These movements increase the distance between the carboxylates, diminish the electrostatic repulsion, and lead to the remarkably high activity of the S113E/N115L mutant.
  Selected figure(s)  
Figure 1.
Fig. 1. Effects n enzyme performance (k,,l/K,.~,o,~trate/Km.~~~~) of the various mutants. The firstletter of he single-letter amino cid code refers to he amino acid at site 113 and the second to the amino acid t site 115.
Figure 3.
Fig. 3. pH profiles: 0, wild type; 0, S113N; , S113D; and U, S113D/NllSL mutants. Disturbance in the V,, profile of the S113D singlemutantisabeninthat of theS113D/N115Ldoublemutant (top), whereas changes in pH do not affect affinity below pH 8.5 (bottom).
  The above figures are reprinted from an Open Access publication published by the Protein Society: Protein Sci (1996, 5, 287-295) copyright 1996.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19365560 A.G.Nackley, S.A.Shabalina, J.E.Lambert, M.S.Conrad, D.G.Gibson, A.N.Spiridonov, S.K.Satterfield, and L.Diatchenko (2009).
Low enzymatic activity haplotypes of the human catechol-O-methyltransferase gene: enrichment for marker SNPs.
  PLoS ONE, 4, e5237.  
17109184 E.S.Haag (2007).
Compensatory vs. pseudocompensatory evolution in molecular and developmental interactions.
  Genetica, 129, 45-55.  
12403824 A.S.Kondrashov, S.Sunyaev, and F.A.Kondrashov (2002).
Dobzhansky-Muller incompatibilities in protein evolution.
  Proc Natl Acad Sci U S A, 99, 14878-14883.  
12218169 M.L.Baker, I.I.Serysheva, S.Sencer, Y.Wu, S.J.Ludtke, W.Jiang, S.L.Hamilton, and W.Chiu (2002).
The skeletal muscle Ca2+ release channel has an oxidoreductase-like domain.
  Proc Natl Acad Sci U S A, 99, 12155-12160.  
11284679 S.A.Doyle, P.T.Beernink, and D.E.Koshland (2001).
Structural basis for a change in substrate specificity: crystal structure of S113E isocitrate dehydrogenase in a complex with isopropylmalate, Mg2+, and NADP.
  Biochemistry, 40, 4234-4241.
PDB code: 1hj6
  11206056 R.Chen, and S.S.Jeong (2000).
Functional prediction: identification of protein orthologs and paralogs.
  Protein Sci, 9, 2344-2353.  
11087384 S.A.Doyle, S.Y.Fung, and D.E.Koshland (2000).
Redesigning the substrate specificity of an enzyme: isocitrate dehydrogenase.
  Biochemistry, 39, 14348-14355.  
9096353 A.M.Dean, and G.B.Golding (1997).
Protein engineering reveals ancient adaptive replacements in isocitrate dehydrogenase.
  Proc Natl Acad Sci U S A, 94, 3104-3109.  
8673602 B.L.Stoddard, A.Dean, and P.A.Bash (1996).
Combining Laue diffraction and molecular dynamics to study enzyme intermediates.
  Nat Struct Biol, 3, 590-595.  
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 code is shown on the right.