Expression, purification, and crystallization of restriction endonuclease PvuII with DNA containing its recognition site.
We have overexpressed the type II restriction endonuclease PvuII (R-PvuII) in E. coli, prepared large amounts of the homogeneous enzyme, and crystallized it with an oligonucleotide carrying a PvuII recognition site. The cocrystals are orthorhombic space group P2(1)2(1)2(1) with cell constants a = 95.8 A, b = 86.3 A, c = 48.5 A, and diffract X-rays to at least 2.7 A. There is a complex of two protein subunits and one oligonucleotide duplex in the asymmetric unit.
Balendiran
K
K
W. M. Keck Structural Biology Laboratory, Cold Spring Harbor Laboratory, New York 11724.
Bonventre
J
J
Knott
R
R
Jack
W
W
Benner
J
J
Schildkraut
I
I
Anderson
J E
JE
D001483:Base Sequence; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli; D015870:Gene Expression; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism
D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; C025630:CAGCTG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific