2HJP

Crystal Structure of Phosphonopyruvate Hydrolase Complex with Phosphonopyruvate and Mg++


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.225 
  • R-Value Work: 0.169 
  • R-Value Observed: 0.175 

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Ligand Structure Quality Assessment 


This is version 1.4 of the entry. See complete history


Literature

Structure and Kinetics of Phosphonopyruvate Hydrolase from Voriovorax sp. Pal2: New Insight into the Divergence of Catalysis within the PEP Mutase/Isocitrate Lyase Superfamily

Chen, C.C.H.Han, Y.Niu, W.Kulakova, A.N.Howard, A.Quinn, J.P.Dunaway-Mariano, D.Herzberg, O.

(2006) Biochemistry 45: 11491-11504

  • DOI: https://doi.org/10.1021/bi061208l
  • Primary Citation of Related Structures:  
    2DUA, 2HJP, 2HRW

  • PubMed Abstract: 

    Phosphonopyruvate (P-pyr) hydrolase (PPH), a member of the phosphoenolpyruvate (PEP) mutase/isocitrate lyase (PEPM/ICL) superfamily, hydrolyzes P-pyr and shares the highest sequence identity and functional similarity with PEPM. Recombinant PPH from Variovorax sp. Pal2 was expressed in Escherichia coli and purified to homogeneity. Analytical gel filtration indicated that the protein exists in solution predominantly as a tetramer. The PPH pH rate profile indicates maximal activity over a broad pH range. The steady-state kinetic constants determined for a rapid equilibrium ordered kinetic mechanism with Mg2+ binding first (Kd = 140 +/- 40 microM), are kcat = 105 +/- 2 s(-1) and P-pyr Km = 5 +/- 1 microM. PEP (slow substrate kcat = 2 x 10(-4) s(-1)), oxalate, and sulfopyruvate are competitive inhibitors with Ki values of 2.0 +/- 0.1 mM, 17 +/- 1 microM, and 210 +/- 10 microM, respectively. Three PPH crystal structures have been determined, that of a ligand-free enzyme, the enzyme bound to Mg2+ and oxalate (inhibitor), and the enzyme bound to Mg2+ and P-pyr (substrate). The complex with the inhibitor was obtained by cocrystallization, whereas that with the substrate was obtained by briefly soaking crystals of the ligand-free enzyme with P-pyr prior to flash cooling. The PPH structure resembles that of the other members of the PEPM/ICL superfamily and is most similar to the functionally related enzyme, PEPM. Each monomer of the dimer of dimers exhibits an (alpha/beta)8 barrel fold with the eighth helix swapped between two molecules of the dimer. Both P-pyr and oxalate are anchored to the active site by Mg2+. The loop capping the active site is disordered in all three structures, in contrast to PEPM, where the equivalent loop adopts an open or disordered conformation in the unbound state but sequesters the inhibitor from solvent in the bound state. Crystal packing may have favored the open conformation of PPH even when the enzyme was cocrystallized with the oxalate inhibitor. Structure alignment of PPH with other superfamily members revealed two pairs of invariant or conservatively replaced residues that anchor the flexible gating loop. The proposed PPH catalytic mechanism is analogous to that of PEPM but includes activation of a water nucleophile with the loop Thr118 residue.


  • Organizational Affiliation

    Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville, Maryland 20850, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Phosphonopyruvate hydrolase290Variovorax sp. Pal2Mutation(s): 0 
Gene Names: pphA
UniProt
Find proteins for Q84G06 (Variovorax sp. (strain Pal2))
Explore Q84G06 
Go to UniProtKB:  Q84G06
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ84G06
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 5 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
PPR
Query on PPR

Download Ideal Coordinates CCD File 
G [auth A]PHOSPHONOPYRUVATE
C3 H5 O6 P
CHDDAVCOAOFSLD-UHFFFAOYSA-N
XYS
Query on XYS

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B [auth A]alpha-D-xylopyranose
C5 H10 O5
SRBFZHDQGSBBOR-LECHCGJUSA-N
CL
Query on CL

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E [auth A],
F [auth A]
CHLORIDE ION
Cl
VEXZGXHMUGYJMC-UHFFFAOYSA-M
MG
Query on MG

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C [auth A]MAGNESIUM ION
Mg
JLVVSXFLKOJNIY-UHFFFAOYSA-N
NA
Query on NA

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D [auth A]SODIUM ION
Na
FKNQFGJONOIPTF-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.225 
  • R-Value Work: 0.169 
  • R-Value Observed: 0.175 
  • Space Group: I 2 2 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 74.89α = 90
b = 79.18β = 90
c = 93.43γ = 90
Software Package:
Software NamePurpose
d*TREKdata processing
CNSrefinement
PDB_EXTRACTdata extraction
d*TREKdata reduction
d*TREKdata scaling
CNSphasing

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2006-10-03
    Type: Initial release
  • Version 1.1: 2008-05-01
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Derived calculations, Source and taxonomy, Version format compliance
  • Version 1.3: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Advisory, Data collection, Derived calculations, Structure summary
  • Version 1.4: 2024-02-14
    Changes: Advisory, Data collection, Database references, Structure summary