Crystallization and preliminary X‐ray diffraction analysis of the homing endonuclease I‐CvuI from Chlorella vulgaris in complex with its target DNA. Issue 2 (1st February 2014) - Redondo, Pilar
Abstract 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 Mg2+ 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.