PDBsum entry 1k89

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protein links
Oxidoreductase PDB id
Protein chain
449 a.a. *
Waters ×238
* Residue conservation analysis
PDB id:
Name: Oxidoreductase
Title: K89l mutant of glutamate dehydrogenase
Structure: Glutamate dehydrogenase. Chain: a. Engineered: yes. Mutation: yes
Source: Clostridium symbiosum. Organism_taxid: 1512. Gene: gdh. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Homo-Hexamer (from PDB file)
2.05Å     R-factor:   0.143    
Authors: T.J.Stillman,A.M.B.Migueis,X.G.Wang,P.J.Baker,K.L.Britton,P. D.W.Rice
Key ref:
T.J.Stillman et al. (1999). Insights into the mechanism of domain closure and substrate specificity of glutamate dehydrogenase from Clostridium symbiosum. J Mol Biol, 285, 875-885. PubMed id: 9878450 DOI: 10.1006/jmbi.1998.2335
05-Jun-98     Release date:   27-Jan-99    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P24295  (DHE2_CLOSY) -  NAD-specific glutamate dehydrogenase
450 a.a.
449 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.  - Glutamate dehydrogenase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: L-glutamate + H2O + NAD+ = 2-oxoglutarate + NH3 + NADH
+ H(2)O
+ NAD(+)
= 2-oxoglutarate
+ NH(3)
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     oxidation-reduction process   3 terms 
  Biochemical function     oxidoreductase activity     3 terms  


DOI no: 10.1006/jmbi.1998.2335 J Mol Biol 285:875-885 (1999)
PubMed id: 9878450  
Insights into the mechanism of domain closure and substrate specificity of glutamate dehydrogenase from Clostridium symbiosum.
T.J.Stillman, A.M.Migueis, X.G.Wang, P.J.Baker, K.L.Britton, P.C.Engel, D.W.Rice.
Comparisons of the structures of glutamate dehydrogenase (GluDH) and leucine dehydrogenase (LeuDH) have suggested that two substitutions, deep within the amino acid binding pockets of these homologous enzymes, from hydrophilic residues to hydrophobic ones are critical components of their differential substrate specificity. When one of these residues, K89, which hydrogen-bonds to the gamma-carboxyl group of the substrate l-glutamate in GluDH, was altered by site-directed mutagenesis to a leucine residue, the mutant enzyme showed increased substrate activity for methionine and norleucine but negligible activity with either glutamate or leucine. In order to understand the molecular basis of this shift in specificity we have determined the crystal structure of the K89L mutant of GluDH from Clostridium symbiosum. Analysis of the structure suggests that further subtle differences in the binding pocket prevent the mutant from using a branched hydrophobic substrate but permit the straight-chain amino acids to be used as substrates.The three-dimensional crystal structure of the GluDH from C. symbiosum has been previously determined in two distinct forms in the presence and absence of its substrate glutamate. A comparison of these two structures has revealed that the enzyme can adopt different conformations by flexing about the cleft between its two domains, providing a motion which is critical for orienting the partners involved in the hydride transfer reaction. It has previously been proposed that this conformational change is triggered by substrate binding. However, analysis of the K89L mutant shows that it adopts an almost identical conformation with that of the wild-type enzyme in the presence of substrate. Comparison of the mutant structure with both the wild-type open and closed forms has enabled us to separate conformational changes associated with substrate binding and domain motion and suggests that the domain closure may well be a property of the wild-type enzyme even in the absence of substrate.
  Selected figure(s)  
Figure 1.
Figure 1. (a) A stereoview of a superposition of the active site residues in the closed form of GluDH (black) with those of the K89L mutant (grey) produced by the MIDAS display program [Ferrin et al 1988]. The glutamate substrate of GluDH is shown (Glu 501) together with the mutated residue (K89L), and atoms within hydrogen-bonding distances are linked with broken lines. (b), (c) and (d) Three stereo representations of the electron density in the active sites of the K89L mutant and the closed and open forms of wild-type GluDH, respectively, produced by the program BOBSCRIPT [Esnouf 1997]. The electron density associated with the substrate can be clearly seen in (c).
Figure 4.
Figure 4. A stereoview (MIDAS) of a superposition of the active-site residues of both the open (white) and closed (black) forms of GluDH. The main movements within the active site on domain closure are to residues Lys113, Val377, Leu378 and Ser380.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (1999, 285, 875-885) copyright 1999.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19966418 I.G.Shabalin, E.V.Filippova, K.M.Polyakov, E.G.Sadykhov, T.N.Safonova, T.V.Tikhonova, V.I.Tishkov, and V.O.Popov (2009).
Structures of the apo and holo forms of formate dehydrogenase from the bacterium Moraxella sp. C-1: towards understanding the mechanism of the closure of the interdomain cleft.
  Acta Crystallogr D Biol Crystallogr, 65, 1315-1325.
PDB codes: 2gsd 3fn4
19425107 M.A.Sharkey, and P.C.Engel (2009).
Modular coenzyme specificity: a domain-swopped chimera of glutamate dehydrogenase.
  Proteins, 77, 268-278.  
18261912 L.Swint-Kruse, and H.F.Fisher (2008).
Enzymatic reaction sequences as coupled multiple traces on a multidimensional landscape.
  Trends Biochem Sci, 33, 104-112.  
17229734 A.Ciulli, D.Y.Chirgadze, A.G.Smith, T.L.Blundell, and C.Abell (2007).
Crystal structure of Escherichia coli ketopantoate reductase in a ternary complex with NADP+ and pantoate bound: substrate recognition, conformational change, and cooperativity.
  J Biol Chem, 282, 8487-8497.
PDB code: 2ofp
16244435 M.I.Khan, K.Ito, H.Kim, H.Ashida, T.Ishikawa, H.Shibata, and Y.Sawa (2005).
Molecular properties and enhancement of thermostability by random mutagenesis of glutamate dehydrogenase from Bacillus subtilis.
  Biosci Biotechnol Biochem, 69, 1861-1870.  
14622249 S.Y.Seah, K.L.Britton, D.W.Rice, Y.Asano, and P.C.Engel (2003).
Kinetic analysis of phenylalanine dehydrogenase mutants designed for aliphatic amino acid dehydrogenase activity with guidance from homology-based modelling.
  Eur J Biochem, 270, 4628-4634.  
12193607 H.Y.Yoon, E.H.Cho, H.Y.Kwon, S.Y.Choi, and S.W.Cho (2002).
Importance of glutamate 279 for the coenzyme binding of human glutamate dehydrogenase.
  J Biol Chem, 277, 41448-41454.  
11258921 M.Nakasako, T.Fujisawa, S.Adachi, T.Kudo, and S.Higuchi (2001).
Large-scale domain movements and hydration structure changes in the active-site cleft of unligated glutamate dehydrogenase from Thermococcus profundus studied by cryogenic X-ray crystal structure analysis and small-angle X-ray scattering.
  Biochemistry, 40, 3069-3079.
PDB code: 1euz
10801498 S.Suresh, S.Turley, F.R.Opperdoes, P.A.Michels, and W.G.Hol (2000).
A potential target enzyme for trypanocidal drugs revealed by the crystal structure of NAD-dependent glycerol-3-phosphate dehydrogenase from Leishmania mexicana.
  Structure, 8, 541-552.
PDB codes: 1evy 1evz
10477256 Y.Xu, G.Bhargava, H.Wu, G.Loeber, and L.Tong (1999).
Crystal structure of human mitochondrial NAD(P)+-dependent malic enzyme: a new class of oxidative decarboxylases.
  Structure, 7, R877-R889.  
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.