View source for 2xe0


==Molecular basis of engineered meganuclease targeting of the endogenous human RAG1 locus==
<StructureSection load='2xe0' size='340' side='right'caption='[[2xe0]], [[Resolution|resolution]] 2.31&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[2xe0]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Chlamydomonas_reinhardtii Chlamydomonas reinhardtii] and [https://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2XE0 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=2XE0 FirstGlance]. <br>
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.31&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ACT:ACETATE+ION'>ACT</scene>, <scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=PGO:S-1,2-PROPANEDIOL'>PGO</scene></td></tr>
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=2xe0 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=2xe0 OCA], [https://pdbe.org/2xe0 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=2xe0 RCSB], [https://www.ebi.ac.uk/pdbsum/2xe0 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=2xe0 ProSAT]</span></td></tr>
</table>
== Evolutionary Conservation ==
[[Image:Consurf_key_small.gif|200px|right]]
Check<jmol>
  <jmolCheckbox>
    <scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/xe/2xe0_consurf.spt"</scriptWhenChecked>
    <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked>
    <text>to colour the structure by Evolutionary Conservation</text>
  </jmolCheckbox>
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=2xe0 ConSurf].
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== Publication Abstract from PubMed ==
Homing endonucleases recognize long target DNA sequences generating an accurate double-strand break that promotes gene targeting through homologous recombination. We have modified the homodimeric I-CreI endonuclease through protein engineering to target a specific DNA sequence within the human RAG1 gene. Mutations in RAG1 produce severe combined immunodeficiency (SCID), a monogenic disease leading to defective immune response in the individuals, leaving them vulnerable to infectious diseases. The structures of two engineered heterodimeric variants and one single-chain variant of I-CreI, in complex with a 24-bp oligonucleotide of the human RAG1 gene sequence, show how the DNA binding is achieved through interactions in the major groove. In addition, the introduction of the G19S mutation in the neighborhood of the catalytic site lowers the reaction energy barrier for DNA cleavage without compromising DNA recognition. Gene-targeting experiments in human cell lines show that the designed single-chain molecule preserves its in vivo activity with higher specificity, further enhanced by the G19S mutation. This is the first time that an engineered meganuclease variant targets the human RAG1 locus by stimulating homologous recombination in human cell lines up to 265 bp away from the cleavage site. Our analysis illustrates the key features for a la carte procedure in protein-DNA recognition design, opening new possibilities for SCID patients whose illness can be treated ex vivo.

Molecular basis of engineered meganuclease targeting of the endogenous human RAG1 locus.,Munoz IG, Prieto J, Subramanian S, Coloma J, Redondo P, Villate M, Merino N, Marenchino M, D'Abramo M, Gervasio FL, Grizot S, Daboussi F, Smith J, Chion-Sotinel I, Paques F, Duchateau P, Alibes A, Stricher F, Serrano L, Blanco FJ, Montoya G Nucleic Acids Res. 2010 Sep 16. PMID:20846960<ref>PMID:20846960</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
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<div class="pdbe-citations 2xe0" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Chlamydomonas reinhardtii]]
[[Category: Homo sapiens]]
[[Category: Large Structures]]
[[Category: Alibes A]]
[[Category: Blanco FJ]]
[[Category: Chion-Sotine I]]
[[Category: Coloma J]]
[[Category: D'Abramo M]]
[[Category: Daboussi F]]
[[Category: Duchateau P]]
[[Category: Gervasio FL]]
[[Category: Grizot S]]
[[Category: Marenchino M]]
[[Category: Merino N]]
[[Category: Montoya G]]
[[Category: Munoz IG]]
[[Category: Paques F]]
[[Category: Prieto J]]
[[Category: Redondo P]]
[[Category: Serrano L]]
[[Category: Smith J]]
[[Category: Stricher F]]
[[Category: Subramanian S]]
[[Category: Villate M]]