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PDBsum entry 1e0t

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protein ligands Protein-protein interface(s) links
Phosphotransferase PDB id
1e0t
Jmol PyMol
Contents
Protein chains
446 a.a. *
Ligands
SO4 ×4
Waters ×126
* Residue conservation analysis
PDB id:
1e0t
Name: Phosphotransferase
Title: R292d mutant of e. Coli pyruvate kinase
Structure: Pyruvate kinase. Chain: a, b, c, d. Synonym: pk. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 511693. Strain: bl21. Expressed in: escherichia coli. Expression_system_taxid: 511693.
Biol. unit: Homo-Tetramer (from PDB file)
Resolution:
1.8Å     R-factor:   0.246     R-free:   0.315
Authors: R.Fortin,A.Mattevi
Key ref:
G.Valentini et al. (2000). The allosteric regulation of pyruvate kinase. J Biol Chem, 275, 18145-18152. PubMed id: 10751408 DOI: 10.1074/jbc.M001870200
Date:
10-Apr-00     Release date:   11-Apr-00    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P0AD61  (KPYK1_ECOLI) -  Pyruvate kinase I
Seq:
Struc:
470 a.a.
446 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.2.7.1.40  - Pyruvate kinase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: ATP + pyruvate = ADP + phosphoenolpyruvate
ATP
+ pyruvate
= ADP
+ phosphoenolpyruvate
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   2 terms 
  Biological process     metabolic process   5 terms 
  Biochemical function     catalytic activity     10 terms  

 

 
    reference    
 
 
DOI no: 10.1074/jbc.M001870200 J Biol Chem 275:18145-18152 (2000)
PubMed id: 10751408  
 
 
The allosteric regulation of pyruvate kinase.
G.Valentini, L.Chiarelli, R.Fortin, M.L.Speranza, A.Galizzi, A.Mattevi.
 
  ABSTRACT  
 
Pyruvate kinase (PK) is critical for the regulation of the glycolytic pathway. The regulatory properties of Escherichia coli were investigated by mutating six charged residues involved in interdomain salt bridges (Arg(271), Arg(292), Asp(297), and Lys(413)) and in the binding of the allosteric activator (Lys(382) and Arg(431)). Arg(271) and Lys(413) are located at the interface between A and C domains within one subunit. The R271L and K413Q mutant enzymes exhibit altered kinetic properties. In K413Q, there is partial enzyme activation, whereas R271L is characterized by a bias toward the T-state in the allosteric equilibrium. In the T-state, Arg(292) and Asp(297) form an intersubunit salt bridge. The mutants R292D and D297R are totally inactive. The crystal structure of R292D reveals that the mutant enzyme retains the T-state quaternary structure. However, the mutation induces a reorganization of the interface with the creation of a network of interactions similar to that observed in the crystal structures of R-state yeast and M1 PK proteins. Furthermore, in the R292D structure, two loops that are part of the active site are disordered. The K382Q and R431E mutations were designed to probe the binding site for fructose 1, 6-bisphosphate, the allosteric activator. R431E exhibits only slight changes in the regulatory properties. Conversely, K382Q displays a highly altered responsiveness to the activator, suggesting that Lys(382) is involved in both activator binding and allosteric transition mechanism. Taken together, these results support the notion that domain interfaces are critical for the allosteric transition. They couple changes in the tertiary and quaternary structures to alterations in the geometry of the fructose 1, 6-bisphosphate and substrate binding sites. These site-directed mutagenesis data are discussed in the light of the molecular basis for the hereditary nonspherocytic hemolytic anemia, which is caused by mutations in human erythrocyte PK gene.
 
  Selected figure(s)  
 
Figure 3.
Fig. 3. The conformation of residues neighboring the amino acid at position 271 in the wild-type (A) and R271L mutant (B). The contact distances indicated by dashed lines are in angstroms (Å).
Figure 5.
Fig. 5. View of the A/A' interface and of the A domain ( / )[8] barrels in the R292D mutant. Residues 281-290 and 314-321 are connected by dashed lines, because they are at the border of loops 7 and 8, which are disordered. The orientation is approximately the same as in Fig. 4.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2000, 275, 18145-18152) copyright 2000.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19801387 J.P.Schmitz, N.A.van Riel, K.Nicolay, P.A.Hilbers, and J.A.Jeneson (2010).
Silencing of glycolysis in muscle: experimental observation and numerical analysis.
  Exp Physiol, 95, 380-397.  
20856875 R.Bakszt, A.Wernimont, A.Allali-Hassani, M.W.Mok, T.Hills, R.Hui, and J.C.Pizarro (2010).
The crystal structure of Toxoplasma gondii pyruvate kinase 1.
  PLoS One, 5, e12736.
PDB codes: 3eoe 3gg8
20857498 V.Gupta, and R.N.Bamezai (2010).
Human pyruvate kinase M2: a multifunctional protein.
  Protein Sci, 19, 2031-2044.  
19265196 K.Akhtar, V.Gupta, A.Koul, N.Alam, R.Bhat, and R.N.Bamezai (2009).
Differential behavior of missense mutations in the intersubunit contact domain of the human pyruvate kinase M2 isozyme.
  J Biol Chem, 284, 11971-11981.  
19085939 R.van Wijk, E.G.Huizinga, A.C.van Wesel, B.A.van Oirschot, M.A.Hadders, and W.W.van Solinge (2009).
Fifteen novel mutations in PKLR associated with pyruvate kinase (PK) deficiency: structural implications of amino acid substitutions in PK.
  Hum Mutat, 30, 446-453.  
18511452 K.Suzuki, S.Ito, A.Shimizu-Ibuka, and H.Sakai (2008).
Crystal structure of pyruvate kinase from Geobacillus stearothermophilus.
  J Biochem, 144, 305-312.
PDB code: 2e28
18326043 T.Saito, M.Nishi, M.I.Lim, B.Wu, T.Maeda, H.Hashimoto, T.Takeuchi, D.S.Roos, and T.Asai (2008).
A novel GDP-dependent pyruvate kinase isozyme from Toxoplasma gondii localizes to both the apicoplast and the mitochondrion.
  J Biol Chem, 283, 14041-14052.  
17360088 A.Zanella, E.Fermo, P.Bianchi, L.R.Chiarelli, and G.Valentini (2007).
Pyruvate kinase deficiency: the genotype-phenotype association.
  Blood Rev, 21, 217-231.  
17892448 S.Baud, S.Wuillème, B.Dubreucq, A.de Almeida, C.Vuagnat, L.Lepiniec, M.Miquel, and C.Rochat (2007).
Function of plastidial pyruvate kinases in seeds of Arabidopsis thaliana.
  Plant J, 52, 405-419.  
18027374 S.S.Kharalkar, G.S.Joshi, F.N.Musayev, M.Fornabaio, D.J.Abraham, and M.K.Safo (2007).
Identification of novel allosteric regulators of human-erythrocyte pyruvate kinase.
  Chem Biodivers, 4, 2603-2617.  
16879645 H.Imanaka, A.Yamatsu, T.Fukui, H.Atomi, and T.Imanaka (2006).
Phosphoenolpyruvate synthase plays an essential role for glycolysis in the modified Embden-Meyerhof pathway in Thermococcus kodakarensis.
  Mol Microbiol, 61, 898-909.  
15982340 A.Zanella, E.Fermo, P.Bianchi, and G.Valentini (2005).
Red cell pyruvate kinase deficiency: molecular and clinical aspects.
  Br J Haematol, 130, 11-25.  
15614759 F.Glaser, Y.Rosenberg, A.Kessel, T.Pupko, and N.Ben-Tal (2005).
The ConSurf-HSSP database: the mapping of evolutionary conservation among homologs onto PDB structures.
  Proteins, 58, 610-617.  
  16511150 K.Suzuki, S.Ito, A.Shimizu-Ibuka, and H.Sakai (2005).
Crystallization and preliminary X-ray analysis of pyruvate kinase from Bacillus stearothermophilus.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 61, 759-761.  
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.

 

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