1ZM7

Crystal structure of D. melanogaster deoxyribonucleoside kinase mutant N64D in complex with dTTP


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.237 
  • R-Value Work: 0.212 
  • R-Value Observed: 0.213 

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This is version 1.5 of the entry. See complete history


Literature

Structural basis for the changed substrate specificity of Drosophila melanogaster deoxyribonucleoside kinase mutant N64D.

Welin, M.Skovgaard, T.Knecht, W.Zhu, C.Berenstein, D.Munch-Petersen, B.Piskur, J.Eklund, H.

(2005) FEBS J 272: 3733-3742

  • DOI: https://doi.org/10.1111/j.1742-4658.2005.04803.x
  • Primary Citation of Related Structures:  
    1ZM7, 1ZMX

  • PubMed Abstract: 

    The Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK) double mutant N45D/N64D was identified during a previous directed evolution study. This mutant enzyme had a decreased activity towards the natural substrates and decreased feedback inhibition with dTTP, whereas the activity with 3'-modified nucleoside analogs like 3'-azidothymidine (AZT) was nearly unchanged. Here, we identify the mutation N64D as being responsible for these changes. Furthermore, we crystallized the mutant enzyme in the presence of one of its substrates, thymidine, and the feedback inhibitor, dTTP. The introduction of the charged Asp residue appears to destabilize the LID region (residues 167-176) of the enzyme by electrostatic repulsion and no hydrogen bond to the 3'-OH is made in the substrate complex by Glu172 of the LID region. This provides a binding space for more bulky 3'-substituents like the azido group in AZT but influences negatively the interactions between Dm-dNK, substrates and feedback inhibitors based on deoxyribose. The detailed picture of the structure-function relationship provides an improved background for future development of novel mutant suicide genes for Dm-dNK-mediated gene therapy.


  • Organizational Affiliation

    Department of Molecular Biology, Swedish University of Agricultural Sciences, Biomedical Center, Uppsala, Sweden.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Deoxynucleoside kinase
A, B, C, D
230Drosophila melanogasterMutation(s): 1 
Gene Names: dnk
EC: 2.7.1.145
UniProt
Find proteins for Q9XZT6 (Drosophila melanogaster)
Explore Q9XZT6 
Go to UniProtKB:  Q9XZT6
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ9XZT6
Sequence Annotations
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  • Reference Sequence
Small Molecules
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.237 
  • R-Value Work: 0.212 
  • R-Value Observed: 0.213 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 67.04α = 90
b = 119.27β = 92.59
c = 68.388γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
MOSFLMdata reduction
CCP4data 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: 2005-05-17
    Type: Initial release
  • Version 1.1: 2008-04-30
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 1.3: 2017-10-11
    Changes: Refinement description
  • Version 1.4: 2021-10-20
    Changes: Database references, Derived calculations
  • Version 1.5: 2023-08-23
    Changes: Data collection, Refinement description