1zmx: Difference between revisions
New page: left|200px<br /><applet load="1zmx" size="450" color="white" frame="true" align="right" spinBox="true" caption="1zmx, resolution 3.10Å" /> '''Crystal structure of... |
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[[Image:1zmx.gif|left|200px]]<br /><applet load="1zmx" size=" | [[Image:1zmx.gif|left|200px]]<br /><applet load="1zmx" size="350" color="white" frame="true" align="right" spinBox="true" | ||
caption="1zmx, resolution 3.10Å" /> | caption="1zmx, resolution 3.10Å" /> | ||
'''Crystal structure of D. melanogaster deoxyribonucleoside kinase N64D mutant in complex with thymidine'''<br /> | '''Crystal structure of D. melanogaster deoxyribonucleoside kinase N64D mutant in complex with thymidine'''<br /> | ||
==Overview== | ==Overview== | ||
The Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK) double | 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. | ||
==About this Structure== | ==About this Structure== | ||
1ZMX is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Drosophila_melanogaster Drosophila melanogaster] with SO4 and THM as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Deoxynucleoside_kinase Deoxynucleoside kinase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.1.145 2.7.1.145] Full crystallographic information is available from [http:// | 1ZMX is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Drosophila_melanogaster Drosophila melanogaster] with <scene name='pdbligand=SO4:'>SO4</scene> and <scene name='pdbligand=THM:'>THM</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Deoxynucleoside_kinase Deoxynucleoside kinase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.1.145 2.7.1.145] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1ZMX OCA]. | ||
==Reference== | ==Reference== | ||
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[[Category: n64d mutant]] | [[Category: n64d mutant]] | ||
''Page seeded by [http:// | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:17:10 2008'' | ||
Revision as of 17:17, 21 February 2008
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Crystal structure of D. melanogaster deoxyribonucleoside kinase N64D mutant in complex with thymidine
OverviewOverview
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.
About this StructureAbout this Structure
1ZMX is a Single protein structure of sequence from Drosophila melanogaster with and as ligands. Active as Deoxynucleoside kinase, with EC number 2.7.1.145 Full crystallographic information is available from OCA.
ReferenceReference
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, FEBS J. 2005 Jul;272(14):3733-42. PMID:16008571
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