2o7m
From Proteopedia
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'''The C-terminal loop of the homing endonuclease I-CreI is essential for DNA binding and cleavage. Identification of a novel site for specificity engineering in the I-CreI scaffold''' | '''The C-terminal loop of the homing endonuclease I-CreI is essential for DNA binding and cleavage. Identification of a novel site for specificity engineering in the I-CreI scaffold''' | ||
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[[Category: Prieto, J.]] | [[Category: Prieto, J.]] | ||
[[Category: Redondo, P.]] | [[Category: Redondo, P.]] | ||
| - | [[Category: | + | [[Category: Dna]] |
| - | [[Category: | + | [[Category: Homing endonuclease]] |
| - | [[Category: | + | [[Category: Hydrolase]] |
| - | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun May 4 10:25:21 2008'' | |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | |
Revision as of 07:25, 4 May 2008
The C-terminal loop of the homing endonuclease I-CreI is essential for DNA binding and cleavage. Identification of a novel site for specificity engineering in the I-CreI scaffold
Overview
Meganucleases are sequence-specific endonucleases with large cleavage sites that can be used to induce efficient homologous gene targeting in cultured cells and plants. These enzymes open novel perspectives for genome engineering in a wide range of fields, including gene therapy. A new crystal structure of the I-CreI dimer without DNA has allowed the comparison with the DNA-bound protein. The C-terminal loop displays a different conformation, which suggests its implication in DNA binding. A site-directed mutagenesis study in this region demonstrates that whereas the C-terminal helix is negligible for DNA binding, the final C-terminal loop is essential in DNA binding and cleavage. We have identified two regions that comprise the Ser138-Lys139 and Lys142-Thr143 pairs whose double mutation affect DNA binding in vitro and abolish cleavage in vivo. However, the mutation of only one residue in these sites allows DNA binding in vitro and cleavage in vivo. These findings demonstrate that the C-terminal loop of I-CreI endonuclease plays a fundamental role in its catalytic mechanism and suggest this novel site as a region to take into account for engineering new endonucleases with tailored specificity.
About this Structure
2O7M is a Single protein structure of sequence from Chlamydomonas reinhardtii. Full crystallographic information is available from OCA.
Reference
The C-terminal loop of the homing endonuclease I-CreI is essential for site recognition, DNA binding and cleavage., Prieto J, Redondo P, Padro D, Arnould S, Epinat JC, Paques F, Blanco FJ, Montoya G, Nucleic Acids Res. 2007;35(10):3262-71. Epub 2007 Apr 22. PMID:17452357 Page seeded by OCA on Sun May 4 10:25:21 2008
