PDBsum entry 2j5t

Go to PDB code: 
protein ligands metals Protein-protein interface(s) links
Transferase PDB id
Protein chains
(+ 1 more) 355 a.a. *
337 a.a. *
GLU ×12
SO4 ×8
_MG ×8
_CL ×20
Waters ×103
* Residue conservation analysis
PDB id:
Name: Transferase
Title: Glutamate 5-kinase from escherichia coli complexed with glutamate
Structure: Glutamate 5-kinase. Chain: a, b, c, d, e, f, g, h. Synonym: gamma-glutamyl kinase, gk. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Strain: dh5alpha. Expressed in: escherichia coli. Expression_system_taxid: 469008.
2.90Å     R-factor:   0.199     R-free:   0.246
Authors: C.Marco-Marin,F.Gil-Ortiz,I.Perez-Arellano,J.Cervera,I.Fita,
Key ref:
C.Marco-Marín et al. (2007). A novel two-domain architecture within the amino acid kinase enzyme family revealed by the crystal structure of Escherichia coli glutamate 5-kinase. J Mol Biol, 367, 1431-1446. PubMed id: 17321544 DOI: 10.1016/j.jmb.2007.01.073
19-Sep-06     Release date:   06-Mar-07    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P0A7B5  (PROB_ECOLI) -  Glutamate 5-kinase
367 a.a.
355 a.a.*
Protein chain
Pfam   ArchSchema ?
P0A7B5  (PROB_ECOLI) -  Glutamate 5-kinase
367 a.a.
337 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chains A, B, C, D, E, F, G, H: E.C.  - Glutamate 5-kinase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

Proline Biosynthesis
      Reaction: ATP + L-glutamate = ADP + L-glutamate 5-phosphate
Bound ligand (Het Group name = GLU)
corresponds exactly
+ L-glutamate 5-phosphate
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     L-proline biosynthetic process   4 terms 
  Biochemical function     nucleotide binding     7 terms  


DOI no: 10.1016/j.jmb.2007.01.073 J Mol Biol 367:1431-1446 (2007)
PubMed id: 17321544  
A novel two-domain architecture within the amino acid kinase enzyme family revealed by the crystal structure of Escherichia coli glutamate 5-kinase.
C.Marco-Marín, F.Gil-Ortiz, I.Pérez-Arellano, J.Cervera, I.Fita, V.Rubio.
Glutamate 5-kinase (G5K) makes the highly unstable product glutamyl 5-phosphate (G5P) in the initial, controlling step of proline/ornithine synthesis, being feedback-inhibited by proline or ornithine, and causing, when defective, clinical hyperammonaemia. We determined two crystal structures of G5K from Escherichia coli, at 2.9 A and 2.5 A resolution, complexed with glutamate and sulphate, or with G5P, sulphate and the proline analogue 5-oxoproline. E. coli G5K presents a novel tetrameric (dimer of dimers) architecture. Each subunit contains a 257 residue AAK domain, typical of acylphosphate-forming enzymes, with characteristic alpha(3)beta(8)alpha(4) sandwich topology. This domain is responsible for catalysis and proline inhibition, and has a crater on the beta sheet C-edge that hosts the active centre and bound 5-oxoproline. Each subunit contains a 93 residue C-terminal PUA domain, typical of RNA-modifying enzymes, which presents the characteristic beta(5)beta(4) sandwich fold and three alpha helices. The AAK and PUA domains of one subunit associate non-canonically in the dimer with the same domains of the other subunit, leaving a negatively charged hole between them that hosts two Mg ions in one crystal, in line with the G5K requirement for free Mg. The tetramer, formed by two dimers interacting exclusively through their AAK domains, is flat and elongated, and has in each face, pericentrically, two exposed active centres in alternate subunits. This would permit the close apposition of two active centres of bacterial glutamate-5-phosphate reductase (the next enzyme in the proline/ornithine-synthesising route), supporting the postulated channelling of G5P. The structures clarify substrate binding and catalysis, justify the high glutamate specificity, explain the effects of known point mutations, and support the binding of proline near glutamate. Proline binding may trigger the movement of a loop that encircles glutamate, and which participates in a hydrogen bond network connecting active centres, which is possibly involved in the cooperativity for glutamate.
  Selected figure(s)  
Figure 1.
Figure 1. Pathway of proline synthesis in microorganisms and plants, and of ornithine synthesis in mammals. Enzymes are enclosed in grey boxes. Feed-back inhibition of microbial and plant G5Ks by proline and of animal G5Ks by ornithine is indicated with broken arrows. The dotted arrow indicates the spontaneous cyclization of G5P to 5-oxoproline that is an abortive side-reaction.
Figure 9.
Figure 9. Possible interaction between the glutamate 5-kinase (G5K) and the glutamyl 5-phosphate reductase (G5PR). Surface representation of two perpendicular views of the G5K tetramer, with the substrates in space-filling representation. A dimer of the G5PR from T. maritima is shown (ribbon representation) with one subunit (orange) presenting the open conformation as observed in the crystal structure of this enzyme in the absence of substrates (PDB 1O20), and with the other subunit (yellow) presenting a closed conformation modelled from class 3 aldehyde dehydrogenase complexed to NAD (PDB 1AD3). The catalytic and NADPH-binding domains of G5PR are identified.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 367, 1431-1446) copyright 2007.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21186348 K.Veeravalli, D.Boyd, B.L.Iverson, J.Beckwith, and G.Georgiou (2011).
Laboratory evolution of glutathione biosynthesis reveals natural compensatory pathways.
  Nat Chem Biol, 7, 101-105.  
  20091669 I.Pérez-Arellano, F.Carmona-Alvarez, A.I.Martínez, J.Rodríguez-Díaz, and J.Cervera (2010).
Pyrroline-5-carboxylate synthase and proline biosynthesis: from osmotolerance to rare metabolic disease.
  Protein Sci, 19, 372-382.  
20544237 I.Pérez-Arellano, and J.Cervera (2010).
Glutamate kinase from Thermotoga maritima: characterization of a thermophilic enzyme for proline biosynthesis.
  Extremophiles, 14, 409-415.  
  20402538 M.F.Mabanglo, H.L.Schubert, M.Chen, C.P.Hill, and C.D.Poulter (2010).
X-ray structures of isopentenyl phosphate kinase.
  ACS Chem Biol, 5, 517-527.
PDB codes: 3lkk 3ll5 3ll9
  20392112 N.Dellas, and J.P.Noel (2010).
Mutation of archaeal isopentenyl phosphate kinase highlights mechanism and guides phosphorylation of additional isoprenoid monophosphates.
  ACS Chem Biol, 5, 589-601.
PDB codes: 3k4o 3k4y 3k52 3k56
19620997 S.D.Copley (2009).
Evolution of efficient pathways for degradation of anthropogenic chemicals.
  Nat Chem Biol, 5, 559-566.  
19734662 T.Kaino, and H.Takagi (2009).
Proline as a stress protectant in the yeast Saccharomyces cerevisiae: effects of trehalose and PRO1 gene expression on stress tolerance.
  Biosci Biotechnol Biochem, 73, 2131-2135.  
18184660 D.Shi, V.Sagar, Z.Jin, X.Yu, L.Caldovic, H.Morizono, N.M.Allewell, and M.Tuchman (2008).
The crystal structure of N-acetyl-L-glutamate synthase from Neisseria gonorrhoeae provides insights into mechanisms of catalysis and regulation.
  J Biol Chem, 283, 7176-7184.
PDB codes: 2r8v 2r98 3b8g
18802692 H.Takagi (2008).
Proline as a stress protectant in yeast: physiological functions, metabolic regulations, and biotechnological applications.
  Appl Microbiol Biotechnol, 81, 211-223.  
18701452 S.Pakhomova, S.G.Bartlett, A.Augustus, T.Kuzuyama, and M.E.Newcomer (2008).
Crystal Structure of Fosfomycin Resistance Kinase FomA from Streptomyces wedmorensis.
  J Biol Chem, 283, 28518-28526.
PDB codes: 3d40 3d41
17803682 I.Pérez-Arellano, J.Gallego, and J.Cervera (2007).
The PUA domain - a structural and functional overview.
  FEBS J, 274, 4972-4984.  
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.