3BGI

Thiopurine S-Methyltransferase


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.80 Å
  • R-Value Free: 0.221 
  • R-Value Work: 0.187 
  • R-Value Observed: 0.189 

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Ligand Structure Quality Assessment 


This is version 1.3 of the entry. See complete history


Literature

Structural Basis of Substrate Recognition in Thiopurine S-Methyltransferase

Peng, Y.Feng, Q.Wilk, D.Adjei, A.A.Salavaggione, O.E.Weinshilboum, R.M.Yee, V.C.

(2008) Biochemistry 47 (23), 6216 6225, 2008. 10.1021/bi800102x: 6216-6225

  • DOI: https://doi.org/10.1021/bi800102x
  • Primary Citation of Related Structures:  
    3BGD, 3BGI

  • PubMed Abstract: 

    Thiopurine S-methyltransferase (TPMT) modulates the cytotoxic effects of thiopurine prodrugs such as 6-mercaptopurine by methylating them in a reaction using S-adenosyl- l-methionine as the donor. Patients with TPMT variant allozymes exhibit diminished levels of protein and/or enzyme activity and are at risk for thiopurine drug-induced toxicity. We have determined two crystal structures of murine TPMT, as a binary complex with the product S-adenosyl- l-homocysteine and as a ternary complex with S-adenosyl- l-homocysteine and the substrate 6-mercaptopurine, to 1.8 and 2.0 A resolution, respectively. Comparison of the structures reveals that an active site loop becomes ordered upon 6-mercaptopurine binding. The positions of the two ligands are consistent with the expected S N2 reaction mechanism. Arg147 and Arg221, the only polar amino acids near 6-mercaptopurine, are highlighted as possible participants in substrate deprotonation. To probe whether these residues are important for catalysis, point mutants were prepared in the human enzyme. Substitution of Arg152 (Arg147 in murine TPMT) with glutamic acid decreases V max and increases K m for 6-mercaptopurine but not K m for S-adenosyl- l-methionine. Substitution at this position with alanine or histidine and similar substitutions of Arg226 (Arg221 in murine TPMT) result in no effect on enzyme activity. The double mutant Arg152Ala/Arg226Ala exhibits a decreased V max and increased K m for 6-mercaptopurine. These observations suggest that either Arg152 or Arg226 may participate in some fashion in the TPMT reaction, with one residue compensating when the other is altered, and that Arg152 may interact with substrate more directly than Arg226, consistent with observations in the murine TPMT crystal structure.


  • Organizational Affiliation

    Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Thiopurine S-methyltransferase
A, B
260Mus musculusMutation(s): 0 
Gene Names: Tpmt
EC: 2.1.1.67
UniProt
Find proteins for O55060 (Mus musculus)
Explore O55060 
Go to UniProtKB:  O55060
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupO55060
Sequence Annotations
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  • Reference Sequence
Small Molecules
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.80 Å
  • R-Value Free: 0.221 
  • R-Value Work: 0.187 
  • R-Value Observed: 0.189 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 62.908α = 90
b = 65.513β = 115.29
c = 72.897γ = 90
Software Package:
Software NamePurpose
DENZOdata reduction
SCALEPACKdata scaling
REFMACrefinement
PDB_EXTRACTdata extraction
HKL-2000data collection
HKL-2000data reduction

Structure Validation

View Full Validation Report



Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2008-06-10
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Version format compliance
  • Version 1.2: 2017-10-25
    Changes: Refinement description
  • Version 1.3: 2024-02-21
    Changes: Data collection, Database references, Derived calculations