Crystallization and preliminary X-ray diffraction analysis of the homing endonuclease I-CvuI from Chlorella vulgaris in complex with its target DNA
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Homing endonucleases are highly specific DNA-cleaving enzymes that recognize long stretches of DNA. The engineering of these enzymes provides novel instruments for genome modification in a wide range of fields, including gene targeting, by inducing specific double-strand breaks. I-CvuI is a homing endonuclease from the green alga Chlorella vulgaris. This enzyme was purified after overexpression in Escherichia coli. Crystallization experiments of I-CvuI in complex with its DNA target in the presence of Mg(2+) yielded crystals suitable for X-ray diffraction analysis. The crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 62.83, b = 83.56, c = 94.40 Å. The self-rotation function and the Matthews coefficient suggested the presence of one protein-DNA complex per asymmetric unit. The crystals diffracted to a resolution limit of 1.9 Å using synchrotron radiation.
|Journal||Acta crystallographica. Section F, Structural biology communications|
|Issue number||Pt 2|
|Number of pages||4|
|Publication status||Published - Feb 2014|
- Chlorella vulgaris/enzymology, Chromatography, Liquid, Crystallization, Crystallography, X-Ray/methods, DNA/metabolism, Electrophoresis, Polyacrylamide Gel, Endonucleases/chemistry, Protein Conformation