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PDBsum entry 1t36

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protein ligands metals Protein-protein interface(s) links
Ligase PDB id
1t36
Jmol
Contents
Protein chains
1058 a.a. *
379 a.a. *
Ligands
PO4 ×5
ADP ×8
ORN ×4
NET ×4
__U ×4
Metals
__K ×22
_CL ×27
_MN ×12
Waters ×3907
* Residue conservation analysis
PDB id:
1t36
Name: Ligase
Title: Crystal structure of e. Coli carbamoyl phosphate synthetase subunit mutant c248d complexed with uridine 5'-monophosphat
Structure: Carbamoyl-phosphate synthase large chain. Chain: a, c, e, g. Synonym: carbamoyl-phosphate synthetase ammonia chain. Engineered: yes. Carbamoyl-phosphate synthase small chain. Chain: b, d, f, h. Synonym: carbamoyl-phosphate synthetase glutamine chain. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Gene: carb, pyra, b0033. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693. Gene: cara, pyra, b0032, z0037, ecs0035. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Dimer (from PQS)
Resolution:
2.10Å     R-factor:   0.179     R-free:   0.209
Authors: J.B.Thoden,X.Huang,F.M.Raushel,H.M.Holden
Key ref:
J.B.Thoden et al. (2004). Long-range allosteric transitions in carbamoyl phosphate synthetase. Protein Sci, 13, 2398-2405. PubMed id: 15322282 DOI: 10.1110/ps.04822704
Date:
24-Apr-04     Release date:   21-Sep-04    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P00968  (CARB_ECOLI) -  Carbamoyl-phosphate synthase large chain
Seq:
Struc:
 
Seq:
Struc:
 
Seq:
Struc:
1073 a.a.
1058 a.a.
Protein chains
Pfam   ArchSchema ?
P0A6F1  (CARA_ECOLI) -  Carbamoyl-phosphate synthase small chain
Seq:
Struc:
382 a.a.
379 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 Enzyme reactions 
   Enzyme class: Chains A, B, C, D, E, F, G, H: E.C.6.3.5.5  - Carbamoyl-phosphate synthase (glutamine-hydrolyzing).
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

      Pathway:
Pyrimidine Biosynthesis
      Reaction: 2 ATP + L-glutamine + HCO3- + H2O = 2 ADP + phosphate + L-glutamate + carbamoyl phosphate
2 × ATP
+
L-glutamine
Bound ligand (Het Group name = ORN)
matches with 90.00% similarity
+ HCO(3)(-)
+ H(2)O
=
2 × ADP
Bound ligand (Het Group name = ADP)
corresponds exactly
+
phosphate
Bound ligand (Het Group name = PO4)
corresponds exactly
+ L-glutamate
+ carbamoyl phosphate
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   3 terms 
  Biological process     metabolic process   13 terms 
  Biochemical function     catalytic activity     9 terms  

 

 
    reference    
 
 
DOI no: 10.1110/ps.04822704 Protein Sci 13:2398-2405 (2004)
PubMed id: 15322282  
 
 
Long-range allosteric transitions in carbamoyl phosphate synthetase.
J.B.Thoden, X.Huang, J.Kim, F.M.Raushel, H.M.Holden.
 
  ABSTRACT  
 
Carbamoyl phosphate synthetase plays a key role in both pyrimidine and arginine biosynthesis by catalyzing the production of carbamoyl phosphate from one molecule of bicarbonate, two molecules of MgATP, and one molecule of glutamine. The enzyme from Escherichia coli consists of two polypeptide chains referred to as the small and large subunits, which contain a total of three separate active sites that are connected by an intramolecular tunnel. The small subunit harbors one of these active sites and is responsible for the hydrolysis of glutamine to glutamate and ammonia. The large subunit binds the two required molecules of MgATP and is involved in assembling the final product. Compounds such as L-ornithine, UMP, and IMP allosterically regulate the enzyme. Here, we report the three-dimensional structure of a site-directed mutant protein of carbamoyl phosphate synthetase from E. coli, where Cys 248 in the small subunit was changed to an aspartate. This residue was targeted for a structural investigation because previous studies demonstrated that the partial glutaminase activity of the C248D mutant protein was increased 40-fold relative to the wild-type enzyme, whereas the formation of carbamoyl phosphate using glutamine as a nitrogen source was completely abolished. Remarkably, although Cys 248 in the small subunit is located at approximately 100 A from the allosteric binding pocket in the large subunit, the electron density map clearly revealed the presence of UMP, although this ligand was never included in the purification or crystallization schemes. The manner in which UMP binds to carbamoyl phosphate synthetase is described.
 
  Selected figure(s)  
 
Figure 3.
Figure 3. Representative electron density. The region near the site of the C248D mutation is shown in A, whereas the electron density corresponding to the bound UMP is displayed in B. The electron density maps shown were calculated with coefficients of the form (F[o] - F[c]) and contoured at 3 .
Figure 4.
Figure 4. Electrostatic interactions between CPS and the IMP or UMP ligands. Schematic representations of the hydrogen bonding patterns between the protein and the IMP and UMP ligands are depicted in A and B, respectively.
 
  The above figures are reprinted by permission from the Protein Society: Protein Sci (2004, 13, 2398-2405) copyright 2004.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21136295 G.Mendoza-Hernández, B.Pérez-Gómez, D.W.Krogmann, E.B.Gutiérrez-Cirlos, and C.Gómez-Lojero (2010).
Interactions of linker proteins with the phycobiliproteins in the phycobilisome substructures of Gloeobacter violaceus.
  Photosynth Res, 106, 247-261.  
18375388 I.Nissim, O.Horyn, I.Nissim, Y.Daikhin, S.L.Wehrli, and M.Yudkoff (2008).
3-isobutylmethylxanthine inhibits hepatic urea synthesis: protection by agmatine.
  J Biol Chem, 283, 15063-15071.  
17209549 J.L.Johnson, J.K.West, A.D.Nelson, and G.D.Reinhart (2007).
Resolving the fluorescence response of Escherichia coli carbamoyl phosphate synthetase: mapping intra- and intersubunit conformational changes.
  Biochemistry, 46, 387-397.  
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.  
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.