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PDBsum entry 1e4e

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protein ligands metals Protein-protein interface(s) links
Ligase PDB id
1e4e
Jmol
Contents
Protein chains
341 a.a. *
Ligands
ADP ×2
PHY ×2
SO4 ×3
GOL ×14
Metals
_MG ×4
Waters ×355
* Residue conservation analysis
PDB id:
1e4e
Name: Ligase
Title: D-alanyl-d-lacate ligase
Structure: Vancomycin/teicoplanin a-type resistance protein chain: a. Synonym: d-alanine--d-lactate ligase, vana ligase. Engineered: yes. Vancomycin/teicoplanin a-type resistance protein chain: b. Synonym: d-alanine--d-lactate ligase, vana ligase. Engineered: yes
Source: Enterococcus faecium. Organism_taxid: 1352. Strain: bm41417. Cellular_location: cytoplasm. Gene: vana. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_taxid: 562
Biol. unit: Tetramer (from PDB file)
Resolution:
2.50Å     R-factor:   0.183     R-free:   0.257
Authors: D.I.Roper
Key ref:
D.I.Roper et al. (2000). The molecular basis of vancomycin resistance in clinically relevant Enterococci: crystal structure of D-alanyl-D-lactate ligase (VanA). Proc Natl Acad Sci U S A, 97, 8921-8925. PubMed id: 10908650 DOI: 10.1073/pnas.150116497
Date:
03-Jul-00     Release date:   28-Jun-01    
PROCHECK
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 Headers
 References

Protein chains
Pfam   ArchSchema ?
P25051  (VANA_ENTFC) -  Vancomycin/teicoplanin A-type resistance protein VanA
Seq:
Struc:
343 a.a.
341 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 4 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.6.1.2.1  - D-alanine--(R)-lactate ligase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: D-alanine + (R)-lactate + ATP = D-alanyl-(R)-lactate + ADP + phosphate
D-alanine
Bound ligand (Het Group name = GOL)
matches with 71.43% similarity
+ (R)-lactate
+ ATP
= D-alanyl-(R)-lactate
+
ADP
Bound ligand (Het Group name = ADP)
corresponds exactly
+ phosphate
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   3 terms 
  Biological process     cell wall organization   4 terms 
  Biochemical function     catalytic activity     8 terms  

 

 
    reference    
 
 
DOI no: 10.1073/pnas.150116497 Proc Natl Acad Sci U S A 97:8921-8925 (2000)
PubMed id: 10908650  
 
 
The molecular basis of vancomycin resistance in clinically relevant Enterococci: crystal structure of D-alanyl-D-lactate ligase (VanA).
D.I.Roper, T.Huyton, A.Vagin, G.Dodson.
 
  ABSTRACT  
 
d-alanine-d-lactate ligase from Enterococcus faecium BM4147 is directly responsible for the biosynthesis of alternate cell-wall precursors in bacteria, which are resistant to the glycopeptide antibiotic vancomycin. The crystal structure has been determined with data extending to 2.5-A resolution. This structure shows that the active site has unexpected interactions and is distinct from previous models for d-alanyl-d-lactate ligase mechanistic studies. It appears that the preference of the enzyme for lactate as a ligand over d-alanine could be mediated by electrostatic effects and/or a hydrogen-bonding network, which principally involve His-244. The structure of d-alanyl-d-lactate ligase provides a revised interpretation of the molecular events that lead to vancomycin resistance.
 
  Selected figure(s)  
 
Figure 2.
Fig. 2. Schematic diagram of the active site of VanA from the structure showing the interaction of various water molecules in addition to amino acid side chains. The protein backbone of the residues in the region of His-244 and Tyr-315 is shown as a solid black line. Hydrogen bond distances between Tyr-315, His-244, and the second subsite carboxyl oxygen of the transition state inhibitor are 2.77 Å and 2.74 Å, respectively. Two phenylalanine residues (F169 and F294) form stacking interactions with the adenine nucleotide rings. Several water molecules in the active site form interactions with the phosphosphate and magnesium atoms as well as amino acid side chains. There is an additional hydrogen bond between water 475 and water 371, which is not shown for clarity.
Figure 3.
Fig. 3. (a) Comparison of active site waters in E. facieum VanA, which replace Lys-215 in E. coli DdlB. Magnesium atoms are shown in gray with water molecules in red. The structures of VanA and DdlB were superimposed and the relative positions of waters in VanA and Lys-215 in DdlB compared. Water 475 in VanA takes an equivalent position to the side-chain nitrogen of Lys-215 in DdlB. Hydrogen-bonding distances between adjacent water molecules, magnesium atoms, and phosphate atoms are not shown for clarity. Other water molecules, notably W371, W569, and W574, coordinate with magnesium ions in the active site of VanA. (b) A stereo representation and 2 F[o] F[c] map showing the active-site residues in contact with the phosphinophosphinate transition state intermediate. The map is contoured at 1 by using the final 2.5-Å resolution map. Residues in the immediate vicinity of the transition state intermediate are marked. The two magnesium ions that coordinate with the phosphate ion of the intermediate and the -phosphate of ADP are displayed in gray, and water molecules in this vicinity are displayed in red. Water 475 in VanA takes an equivalent position to the side-chain nitrogen of Lys-215 in DdlB. Other water molecules, notably W371 and W574, coordinate with magnesium ions in the active site of VanA. (c) Stereo diagram of the hydrogen bonding interacts with and in the vicinity of the phosphorylated phosphinate inhibitor in the active site. The Glu-250, Lys-22, Tyr-4315, and His-244 hydrogen-bonding network is shown, making a 2.7-Å hydrogen bond with the carboxylate oxygen of the inhibitor. This carboxylate also hydrogen bonds to a conserved serine (316 in VanA), which is not shown for clarity. The position of His-244 in our structure is such that it cannot make hydrogen-bonding interactions with the Glu-16 and Ser-177, which are structurally conserved in comparison to DdlB. The later two amino acids form important interactions that anchor D-Ala in the first subsite, an analogous situation to that found in DdlB.
 
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19770507 Y.Kitamura, A.Ebihara, Y.Agari, A.Shinkai, K.Hirotsu, and S.Kuramitsu (2009).
Structure of D-alanine-D-alanine ligase from Thermus thermophilus HB8: cumulative conformational change and enzyme-ligand interactions.
  Acta Crystallogr D Biol Crystallogr, 65, 1098-1106.  
18320587 D.Wu, L.Zhang, Y.Kong, J.Du, S.Chen, J.Chen, J.Ding, H.Jiang, and X.Shen (2008).
Enzymatic characterization and crystal structure analysis of the D-alanine-D-alanine ligase from Helicobacter pylori.
  Proteins, 72, 1148-1160.
PDB code: 2pvp
18266853 H.Barreteau, A.Kovac, A.Boniface, M.Sova, S.Gobec, and D.Blanot (2008).
Cytoplasmic steps of peptidoglycan biosynthesis.
  FEMS Microbiol Rev, 32, 168-207.  
16779845 J.H.Lee, Y.Na, H.E.Song, D.Kim, B.H.Park, S.H.Rho, Y.J.Im, M.K.Kim, G.B.Kang, D.S.Lee, and S.H.Eom (2006).
Crystal structure of the apo form of D-alanine: D-alanine ligase (Ddl) from Thermus caldophilus: a basis for the substrate-induced conformational changes.
  Proteins, 64, 1078-1082.
PDB code: 2fb9
17015835 S.Liu, J.S.Chang, J.T.Herberg, M.M.Horng, P.K.Tomich, A.H.Lin, and K.R.Marotti (2006).
Allosteric inhibition of Staphylococcus aureus D-alanine:D-alanine ligase revealed by crystallographic studies.
  Proc Natl Acad Sci U S A, 103, 15178-15183.
PDB codes: 2i80 2i87 2i8c
16699191 X.Li, A.V.Volkov, K.Szalewicz, and P.Coppens (2006).
Interaction energies between glycopeptide antibiotics and substrates in complexes determined by X-ray crystallography: application of a theoretical databank of aspherical atoms and a symmetry-adapted perturbation theory-based set of interatomic potentials.
  Acta Crystallogr D Biol Crystallogr, 62, 639-647.  
16158456 G.E.Besong, J.M.Bostock, W.Stubbings, I.Chopra, D.I.Roper, A.J.Lloyd, C.W.Fishwick, and A.P.Johnson (2005).
A de novo designed inhibitor of D-Ala-D-Ala ligase from E. coli.
  Angew Chem Int Ed Engl, 44, 6403-6406.  
12832755 E.Potterton, P.Briggs, M.Turkenburg, and E.Dodson (2003).
A graphical user interface to the CCP4 program suite.
  Acta Crystallogr D Biol Crystallogr, 59, 1131-1137.  
12390279 B.Henriques Normark, and S.Normark (2002).
Antibiotic tolerance in pneumococci.
  Clin Microbiol Infect, 8, 613-622.  
11807177 J.Pootoolal, J.Neu, and G.D.Wright (2002).
Glycopeptide antibiotic resistance.
  Annu Rev Pharmacol Toxicol, 42, 381-408.  
11751117 O.H.Ambúr, P.E.Reynolds, and C.A.Arias (2002).
D-Ala:D-Ala ligase gene flanking the vanC cluster: evidence for presence of three ligase genes in vancomycin-resistant Enterococcus gallinarum BM4174.
  Antimicrob Agents Chemother, 46, 95.  
11759084 N.Woodford (2001).
Epidemiology of the genetic elements responsible for acquired glycopeptide resistance in enterococci.
  Microb Drug Resist, 7, 229-236.  
11083656 L.M.Dalla Costa, P.E.Reynolds, H.A.Souza, D.C.Souza, M.F.Palepou, and N.Woodford (2000).
Characterization of a divergent vanD-type resistance element from the first glycopeptide-resistant strain of Enterococcus faecium isolated in Brazil.
  Antimicrob Agents Chemother, 44, 3444-3446.  
10903933 V.L.Healy, L.S.Mullins, X.Li, S.E.Hall, F.M.Raushel, and C.T.Walsh (2000).
D-Ala-D-X ligases: evaluation of D-alanyl phosphate intermediate by MIX, PIX and rapid quench studies.
  Chem Biol, 7, 505-514.  
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 code is shown on the right.