3URQ

Crystal Structure of PTE mutant H254G/H257W/L303T/M317L/I106C/F132I/L271I/K185R/I274N/A80V/R67H with cyclohexyl methylphosphonate inhibitor


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.10 Å
  • R-Value Free: 0.235 
  • R-Value Work: 0.203 

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Ligand Structure Quality Assessment 


This is version 1.3 of the entry. See complete history


Literature

Enzymes for the homeland defense: optimizing phosphotriesterase for the hydrolysis of organophosphate nerve agents.

Tsai, P.C.Fox, N.Bigley, A.N.Harvey, S.P.Barondeau, D.P.Raushel, F.M.

(2012) Biochemistry 51: 6463-6475

  • DOI: https://doi.org/10.1021/bi300811t
  • Primary Citation of Related Structures:  
    3UPM, 3UR2, 3UR5, 3URA, 3URB, 3URN, 3URQ

  • PubMed Abstract: 

    Phosphotriesterase (PTE) from soil bacteria is known for its ability to catalyze the detoxification of organophosphate pesticides and chemical warfare agents. Most of the organophosphate chemical warfare agents are a mixture of two stereoisomers at the phosphorus center, and the S(P)-enantiomers are significantly more toxic than the R(P)-enantiomers. In previous investigations, PTE variants were created through the manipulation of the substrate binding pockets and these mutants were shown to have greater catalytic activities for the detoxification of the more toxic S(P)-enantiomers of nerve agent analogues for GB, GD, GF, VX, and VR than the less toxic R(P)-enantiomers. In this investigation, alternate strategies were employed to discover additional PTE variants with significant improvements in catalytic activities relative to that of the wild-type enzyme. Screening and selection techniques were utilized to isolate PTE variants from randomized libraries and site specific modifications. The catalytic activities of these newly identified PTE variants toward the S(P)-enantiomers of chromophoric analogues of GB, GD, GF, VX, and VR have been improved up to 15000-fold relative to that of the wild-type enzyme. The X-ray crystal structures of the best PTE variants were determined. Characterization of these mutants with the authentic G-type nerve agents has confirmed the expected improvements in catalytic activity against the most toxic enantiomers of GB, GD, and GF. The values of k(cat)/K(m) for the H257Y/L303T (YT) mutant for the hydrolysis of GB, GD, and GF were determined to be 2 × 10(6), 5 × 10(5), and 8 × 10(5) M(-1) s(-1), respectively. The YT mutant is the most proficient enzyme reported thus far for the detoxification of G-type nerve agents. These results support a combinatorial strategy of rational design and directed evolution as a powerful tool for the discovery of more efficient enzymes for the detoxification of organophosphate nerve agents.


  • Organizational Affiliation

    Department of Chemistry, P.O. Box 30012, Texas A&M University, College Station, TX 77842, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Parathion hydrolase
A, B
327Brevundimonas diminutaMutation(s): 11 
Gene Names: opd
EC: 3.1.8.1
UniProt
Find proteins for P0A434 (Brevundimonas diminuta)
Explore P0A434 
Go to UniProtKB:  P0A434
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP0A434
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 3 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
QMP
Query on QMP

Download Ideal Coordinates CCD File 
E [auth A],
K [auth B]
cyclohexyl methylphosphonate
C7 H15 O3 P
SWYGLBJVTQMYSE-UHFFFAOYSA-N
IMD
Query on IMD

Download Ideal Coordinates CCD File 
F [auth A],
G [auth A],
H [auth A],
L [auth B]
IMIDAZOLE
C3 H5 N2
RAXXELZNTBOGNW-UHFFFAOYSA-O
CO
Query on CO

Download Ideal Coordinates CCD File 
C [auth A],
D [auth A],
I [auth B],
J [auth B]
COBALT (II) ION
Co
XLJKHNWPARRRJB-UHFFFAOYSA-N
Modified Residues  1 Unique
IDChains TypeFormula2D DiagramParent
KCX
Query on KCX
A, B
L-PEPTIDE LINKINGC7 H14 N2 O4LYS
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.10 Å
  • R-Value Free: 0.235 
  • R-Value Work: 0.203 
  • Space Group: P 21 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 85.493α = 90
b = 85.278β = 90
c = 87.868γ = 90
Software Package:
Software NamePurpose
HKL-2000data collection
PHASERphasing
CNSrefinement
HKL-2000data reduction
HKL-2000data scaling

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2012-08-01
    Type: Initial release
  • Version 1.1: 2012-10-03
    Changes: Database references
  • Version 1.2: 2023-09-13
    Changes: Data collection, Database references, Derived calculations, Refinement description
  • Version 1.3: 2023-12-06
    Changes: Data collection