spacer
spacer
Go to PDB code: 
protein ligands Protein-protein interface(s) links
Transferase (glutamine amidotransferase) PDB id
1ecg
Jmol
Contents
Protein chains
492 a.a. *
Ligands
ONL ×2
PIN ×6
Waters ×979
* Residue conservation analysis
PDB id:
1ecg
Name: Transferase (glutamine amidotransferase)
Title: Don inactivated escherichia coli glutamine phosphoribosylpyr (prpp) amidotransferase
Structure: Glutamine phosphoribosylpyrophosphate amidotransf chain: a, b. Engineered: yes
Source: Escherichia coli. Organism_taxid: 562. Cell_line: bl21. Gene: purf. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Other_details: t7 phi10 promoter
Biol. unit: Tetramer (from PDB file)
Resolution:
2.30Å     R-factor:   0.147     R-free:   0.229
Authors: J.M.Krahn
Key ref:
J.H.Kim et al. (1996). Structure and function of the glutamine phosphoribosylpyrophosphate amidotransferase glutamine site and communication with the phosphoribosylpyrophosphate site. J Biol Chem, 271, 15549-15557. PubMed id: 8663035 DOI: 10.1074/jbc.271.26.15549
Date:
23-Apr-96     Release date:   08-Nov-96    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P0AG16  (PUR1_ECOLI) -  Amidophosphoribosyltransferase
Seq:
Struc: 		505 a.a.
492 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.2.4.2.14  - Amidophosphoribosyltransferase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: 5-phospho-beta-D-ribosylamine + diphosphate + L-glutamate = L-glutamine + 5-phospho-alpha-D-ribose 1-diphosphate + H2O
5-phospho-beta-D-ribosylamine
 + diphosphate
 +
L-glutamate
Bound ligand (Het Group name = ONL)
matches with 81.82% similarity 		= L-glutamine
 + 5-phospho-alpha-D-ribose 1-diphosphate
 + H(2)O
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   2 terms 
  Biological process     metabolic process   5 terms 
  Biochemical function     transferase activity     4 terms  

 

 
    reference    
 
 
DOI no: 10.1074/jbc.271.26.15549 J Biol Chem 271:15549-15557 (1996)
PubMed id: 8663035  
 
 
Structure and function of the glutamine phosphoribosylpyrophosphate amidotransferase glutamine site and communication with the phosphoribosylpyrophosphate site.
J.H.Kim, J.M.Krahn, D.R.Tomchick, J.L.Smith, H.Zalkin.
 
  ABSTRACT  
 
Glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase from Escherichia coli exhibits a basal PRPP-independent glutaminase activity having a kcat/Km that is 0.3% of fully active enzyme. Binding of PRPP activates the enzyme by a structural change that lowers the Km for glutamine 100-fold and couples glutamine hydrolysis to synthesis of 5-phosphoribosylamine. By analysis of the x-ray structure of the glutamine site containing bound 6-diazo-5-oxonorleucine, a glutamine affinity analog, and by site-directed mutagenesis we have identified residues important for glutamine binding, catalysis, and coupling with PRPP. Tyr74 is a key residue in the coupling between the sites for glutamine in the NH2-terminal domain and PRPP in the COOH-terminal domain. Arg73 and Asp127 have roles in glutamine binding. The x-ray structure indicates that there are no amino acid side chains sufficiently close to Cys1 to participate as a proton acceptor in formation of the thiolate needed for nucleophilic attack on the carboxamide of glutamine, nor as a general acid for amide nitrogen transfer. Based on the x-ray model of the glutamine site and analysis of a mutant enzyme we propose that the free NH2 terminus of Cys1 functions as the proton acceptor and donor. The results indicate that the side chain of Asn101 and the backbone nitrogen of Gly102 function to stabilize a tetrahedral oxyanion resulting from attack of Cys1 on the glutamine carboxamide. Cys1, Arg73, Asn101, Gly102, and Asp127 are conserved in the NH2-terminal domain of a subfamily of amidotransferases that includes asparagine synthetase, glucosamine 6-phosphate synthase, and glutamate synthase, implying a common function in the four enzymes. Tyr74, on the other hand, is conserved only in glutamine PRPP amidotransferase sequences consistent with a specific role in interdomain coupling. The catalytic framework of key glutamine site residues supports the assignment of glutamine PRPP amidotransferase to a recently described Ntn (NH2-terminal nucleophile) hydrolase family of enzymes.
 
  Selected figure(s)  
 
Figure 3.
Fig. 3. Electron density map. Refined model of the DON-labeled active site overlaid with the original unbiased difference electron density map, contoured at 3 . The map was calculated using the native structure with Cys1 S, Asp127 carboxyl, and neighboring water molecules removed. Atoms are colored as follows: peptide C, yellow; DON C, white; N, blue; O, red; S, green. 		Figure 4.
Fig. 4. Interactions of DON in the glutamine site. Schematic representation of the DON-labeled active site illustrating hydrogen bond interactions involving DON and key residues. DON and the Cys1 S-DON C[6] thioether bond are shown in red. Additional hydrogen bonds between DON and water molecules are not shown. Distances between atoms are given in Table III.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (1996, 271, 15549-15557) copyright 1996.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20080736 M.Bokhove, P.N.Jimenez, W.J.Quax, and B.W.Dijkstra (2010).
The quorum-quenching N-acyl homoserine lactone acylase PvdQ is an Ntn-hydrolase with an unusual substrate-binding pocket.
  Proc Natl Acad Sci U S A, 107, 686-691.
PDB codes: 2wyb 2wyc 2wyd 2wye
19670211 R.Koike, A.Kidera, and M.Ota (2009).
Alteration of oligomeric state and domain architecture is essential for functional transformation between transferase and hydrolase with the same scaffold.
  Protein Sci, 18, 2060-2066.  
18712276 Y.Zhang, M.Morar, and S.E.Ealick (2008).
Structural biology of the purine biosynthetic pathway.
  Cell Mol Life Sci, 65, 3699-3724.  
17922651 P.V.Vrzheshch (2007).
Steady-state kinetics of bifunctional enzymes. Taking into account kinetic hierarchy of fast and slow catalytic cycles in a generalized model.
  Biochemistry (Mosc), 72, 936-943.  
17951049 S.Mouilleron, and B.Golinelli-Pimpaneau (2007).
Conformational changes in ammonia-channeling glutamine amidotransferases.
  Curr Opin Struct Biol, 17, 653-664.  
18001139 V.Fresquet, L.Williams, and F.M.Raushel (2007).
Partial randomization of the four sequential amidation reactions catalyzed by cobyric acid synthetase with a single point mutation.
  Biochemistry, 46, 13983-13993.  
17407260 Y.N.Kang, A.Tran, R.H.White, and S.E.Ealick (2007).
A novel function for the N-terminal nucleophile hydrolase fold demonstrated by the structure of an archaeal inosine monophosphate cyclohydrolase.
  Biochemistry, 46, 5050-5062.
PDB codes: 2ntk 2ntl 2ntm
16669783 R.Zrenner, M.Stitt, U.Sonnewald, and R.Boldt (2006).
Pyrimidine and purine biosynthesis and degradation in plants.
  Annu Rev Plant Biol, 57, 805-836.  
15670165 M.Willemoës, A.Mølgaard, E.Johansson, and J.Martinussen (2005).
Lid L11 of the glutamine amidotransferase domain of CTP synthase mediates allosteric GTP activation of glutaminase activity.
  FEBS J, 272, 856-864.  
12764229 B.A.Manjasetty, J.Powlowski, and A.Vrielink (2003).
Crystal structure of a bifunctional aldolase-dehydrogenase: sequestering a reactive and volatile intermediate.
  Proc Natl Acad Sci U S A, 100, 6992-6997.
PDB code: 1nvm
12354108 M.Willemoës, and B.W.Sigurskjold (2002).
Steady-state kinetics of the glutaminase reaction of CTP synthase from Lactococcus lactis. The role of the allosteric activator GTP incoupling between glutamine hydrolysis and CTP synthesis.
  Eur J Biochem, 269, 4772-4779.  
11483578 L.A.Nahum, and M.Riley (2001).
Divergence of function in sequence-related groups of Escherichia coli proteins.
  Genome Res, 11, 1375-1381.  
11395405 X.Huang, H.M.Holden, and F.M.Raushel (2001).
Channeling of substrates and intermediates in enzyme-catalyzed reactions.
  Annu Rev Biochem, 70, 149-180.  
10850988 A.K.Bera, S.Chen, J.L.Smith, and H.Zalkin (2000).
Temperature-dependent function of the glutamine phosphoribosylpyrophosphate amidotransferase ammonia channel and coupling with glycinamide ribonucleotide synthetase in a hyperthermophile.
  J Bacteriol, 182, 3734-3739.  
  10673422 D.Kohls, T.Sulea, E.O.Purisima, R.E.MacKenzie, and A.Vrielink (2000).
The crystal structure of the formiminotransferase domain of formiminotransferase-cyclodeaminase: implications for substrate channeling in a bifunctional enzyme.
  Structure, 8, 35-46.
PDB code: 1qd1
10387030 F.M.Raushel, J.B.Thoden, and H.M.Holden (1999).
The amidotransferase family of enzymes: molecular machines for the production and delivery of ammonia.
  Biochemistry, 38, 7891-7899.  
10090755 H.G.Schnizer, S.K.Boehlein, J.D.Stewart, N.G.Richards, and S.M.Schuster (1999).
Formation and isolation of a covalent intermediate during the glutaminase reaction of a class II amidotransferase.
  Biochemistry, 38, 3677-3682.  
  10049369 S.Li, J.L.Smith, and H.Zalkin (1999).
Mutational analysis of Bacillus subtilis glutamine phosphoribosylpyrophosphate amidotransferase propeptide processing.
  J Bacteriol, 181, 1403-1408.  
10587437 T.M.Larsen, S.K.Boehlein, S.M.Schuster, N.G.Richards, J.B.Thoden, H.M.Holden, and I.Rayment (1999).
Three-dimensional structure of Escherichia coli asparagine synthetase B: a short journey from substrate to product.
  Biochemistry, 38, 16146-16157.
PDB code: 1ct9
10089364 T.M.Weaver, W.Wang, and S.E.Ealick (1999).
Purification, crystallization and preliminary X-ray diffraction data from selenomethionine glycinamide ribonucleotide synthetase.
  Acta Crystallogr D Biol Crystallogr, 55, 518-521.  
  9514258 C.R.Muchmore, J.M.Krahn, J.H.Kim, H.Zalkin, and J.L.Smith (1998).
Crystal structure of glutamine phosphoribosylpyrophosphate amidotransferase from Escherichia coli.
  Protein Sci, 7, 39-51.
PDB codes: 1ecf 1ecj
  9551555 D.R.Tomchick, R.J.Turner, R.L.Switzer, and J.L.Smith (1998).
Adaptation of an enzyme to regulatory function: structure of Bacillus subtilis PyrR, a pyr RNA-binding attenuation protein and uracil phosphoribosyltransferase.
  Structure, 6, 337-350.
PDB codes: 1a3c 1a4x
9914248 J.L.Smith (1998).
Glutamine PRPP amidotransferase: snapshots of an enzyme in action.
  Curr Opin Struct Biol, 8, 686-694.  
9748330 S.K.Boehlein, J.D.Stewart, E.S.Walworth, R.Thirumoorthy, N.G.Richards, and S.M.Schuster (1998).
Kinetic mechanism of Escherichia coli asparagine synthetase B.
  Biochemistry, 37, 13230-13238.  
9843369 W.Wang, T.J.Kappock, J.Stubbe, and S.E.Ealick (1998).
X-ray crystal structure of glycinamide ribonucleotide synthetase from Escherichia coli.
  Biochemistry, 37, 15647-15662.
PDB code: 1gso
9145098 J.A.Brannigan, and G.G.Dodson (1997).
A short cut for the immune system.
  Nat Struct Biol, 4, 334-338.  
9333323 J.M.Krahn, J.H.Kim, M.R.Burns, R.J.Parry, H.Zalkin, and J.L.Smith (1997).
Coupled formation of an amidotransferase interdomain ammonia channel and a phosphoribosyltransferase active site.
  Biochemistry, 36, 11061-11068.
PDB codes: 1ecb 1ecc
9254614 S.K.Boehlein, E.S.Walworth, and S.M.Schuster (1997).
Identification of cysteine-523 in the aspartate binding site of Escherichia coli asparagine synthetase B.
  Biochemistry, 36, 10168-10177.  
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.