Crystallization and preliminary X-ray diffraction analysis of the homing endonuclease I-CvuI from Chlorella vulgaris in complex with its target DNA

TY - JOUR

T1 - Crystallization and preliminary X-ray diffraction analysis of the homing endonuclease I-CvuI from Chlorella vulgaris in complex with its target DNA

AU - Redondo, Pilar

AU - Merino, Nekane

AU - Villate, Maider

AU - Blanco, Francisco J

AU - Montoya, Guillermo

AU - Molina, Rafael

PY - 2014/2

Y1 - 2014/2

N2 - 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.

AB - 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.

KW - Chlorella vulgaris/enzymology

KW - Chromatography, Liquid

KW - Crystallization

KW - Crystallography, X-Ray/methods

KW - DNA/metabolism

KW - Electrophoresis, Polyacrylamide Gel

KW - Endonucleases/chemistry

KW - Protein Conformation

U2 - 10.1107/S2053230X1400065X

DO - 10.1107/S2053230X1400065X

M3 - Journal article

C2 - 24637769

SN - 2053-230X

VL - 70

SP - 256

EP - 259

JO - Acta crystallographica. Section F, Structural biology communications

JF - Acta crystallographica. Section F, Structural biology communications

IS - Pt 2

ER -