spacer
spacer

PDBsum entry 1ur5

Go to PDB code: 
protein ligands metals Protein-protein interface(s) links
Oxidoreductase PDB id
1ur5
Jmol
Contents
Protein chains
299 a.a. *
Ligands
NAD ×2
Metals
_NA ×2
_CD ×6
_CL
Waters ×547
* Residue conservation analysis
PDB id:
1ur5
Name: Oxidoreductase
Title: Stabilization of a tetrameric malate dehydrogenase by introduction of a disulfide bridge at the dimer/dimer interface
Structure: Malate dehydrogenase. Chain: a, c. Engineered: yes. Mutation: yes
Source: Chloroflexus aurantiacus. Organism_taxid: 1108. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Tetramer (from PDB file)
Resolution:
1.75Å     R-factor:   0.190     R-free:   0.223
Authors: A.Bjork,B.Dalhus,D.Mantzilas,V.G.H.Eijsink,R.Sirevag
Key ref:
A.Bjørk et al. (2003). Stabilization of a tetrameric malate dehydrogenase by introduction of a disulfide bridge at the dimer-dimer interface. J Mol Biol, 334, 811-821. PubMed id: 14636605 DOI: 10.1016/j.jmb.2003.10.006
Date:
27-Oct-03     Release date:   05-Nov-03    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P80040  (MDH_CHLAA) -  Malate dehydrogenase
Seq:
Struc:
309 a.a.
299 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.1.1.1.37  - Malate dehydrogenase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

      Pathway:
Citric acid cycle
      Reaction: (S)-malate + NAD+ = oxaloacetate + NADH
(S)-malate
+
NAD(+)
Bound ligand (Het Group name = NAD)
corresponds exactly
= oxaloacetate
+ NADH
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     oxidation-reduction process   5 terms 
  Biochemical function     catalytic activity     4 terms  

 

 
    reference    
 
 
DOI no: 10.1016/j.jmb.2003.10.006 J Mol Biol 334:811-821 (2003)
PubMed id: 14636605  
 
 
Stabilization of a tetrameric malate dehydrogenase by introduction of a disulfide bridge at the dimer-dimer interface.
A.Bjørk, B.Dalhus, D.Mantzilas, V.G.Eijsink, R.Sirevåg.
 
  ABSTRACT  
 
Malate dehydrogenase (MDH) from the moderately thermophilic bacterium Chloroflexus aurantiacus (CaMDH) is a tetrameric enzyme, while MDHs from mesophilic organisms usually are dimers. To investigate the potential contribution of the extra dimer-dimer interface in CaMDH with respect to thermal stability, we have engineered an intersubunit disulfide bridge designed to strengthen dimer-dimer interactions. The resulting mutant (T187C, containing two 187-187 disulfide bridges in the tetramer) showed a 200-fold increase in half-life at 75 degrees C and an increase of 15 deg. C in apparent melting temperature compared to the wild-type. The crystal structure of the mutant (solved at 1.75 A resolution) was essentially identical with that of the wild-type, with the exception of the added inter-dimer disulfide bridge and the loss of an aromatic intra-dimer contact. Remarkably, the mutant and the wild-type had similar temperature optima and activities at their temperature optima, thus providing a clear case of uncoupling of thermal stability and thermoactivity. The results show that tetramerization may contribute to MDH stability to an extent that depends strongly on the number of stabilizing interactions in the dimer-dimer interface.
 
  Selected figure(s)  
 
Figure 1.
Figure 1. Ribbon diagram of homotetrameric malate dehydrogenase from C. aurantiacus. The tetramer consists of two dimers (AB and CD; see the text), Thr187 (side-chain shown as ball-and-sticks) occurs at the dimer-dimer interface. The NAD cofactor bound close to the catalytic center is shown in yellow.
Figure 4.
Figure 4. Temperature-induced denaturation. The curves represent wild-type (0m) T87C[red] (sB) and T187C[ox] ( open ), and were determined by monitoring ellipticity at 222 nm. The protein concentration was 0.1 mg/ml in 10 mM potassium phosphate buffer, pH 7.5. The path length was 0.1 mm.
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2003, 334, 811-821) copyright 2003.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20614086 D.J.Caruana, and S.Howorka (2010).
Biosensors and biofuel cells with engineered proteins.
  Mol Biosyst, 6, 1548-1556.  
19366703 P.I.Toivanen, T.Nieminen, L.Viitanen, A.Alitalo, M.Roschier, S.Jauhiainen, J.E.Markkanen, O.H.Laitinen, T.T.Airenne, T.A.Salminen, M.S.Johnson, K.J.Airenne, and S.Ylä-Herttuala (2009).
Novel vascular endothelial growth factor D variants with increased biological activity.
  J Biol Chem, 284, 16037-16048.  
18260103 E.Goihberg, O.Dym, S.Tel-Or, L.Shimon, F.Frolow, M.Peretz, and Y.Burstein (2008).
Thermal stabilization of the protozoan Entamoeba histolytica alcohol dehydrogenase by a single proline substitution.
  Proteins, 72, 711-719.
PDB codes: 2nvb 2oui
16807887 J.L.Pellequer, and S.W.Chen (2006).
Multi-template approach to modeling engineered disulfide bonds.
  Proteins, 65, 192-202.  
16930136 R.Ladenstein, and B.Ren (2006).
Protein disulfides and protein disulfide oxidoreductases in hyperthermophiles.
  FEBS J, 273, 4170-4185.  
16151092 T.Kaper, B.Talik, T.J.Ettema, H.Bos, M.J.van der Maarel, and L.Dijkhuizen (2005).
Amylomaltase of Pyrobaculum aerophilum IM2 produces thermoreversible starch gels.
  Appl Environ Microbiol, 71, 5098-5106.  
15857780 V.G.Eijsink, S.Gåseidnes, T.V.Borchert, and B.van den Burg (2005).
Directed evolution of enzyme stability.
  Biomol Eng, 22, 21-30.  
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.