Purification and crystallization of Escherichia coli oligoribonuclease.
Oligoribonuclease (Orn) is an essential 3'-to-5' hydrolytic exoribonuclease which degrades short oligoribonucleotides to 5' mononucleotides. Escherichia coli Orn has been crystallized under several different conditions using ammonium sulfate, sodium citrate and sodium acetate as precipitants. Both native and selenomethionine-labeled oligoribonuclease (SeMet-Orn) can be crystallized at room temperature in 1.4-1.55 M sodium citrate. The SeMet-Orn crystals diffract to 2.2 A resolution and belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 70.43, b = 72.87, c = 147.76 A, and two dimers in the asymmetric unit. When grown in the presence of manganese, a second crystal form (Mn-SeMet-Orn) was obtained containing a single dimer per asymmetric unit (P2(1)2(1)2(1); a = 63.74, b = 74.31, c = 74.19 A). Finally, a hexagonal crystal form was obtained using sodium acetate as a precipitant (a = 91.5, b = 91.5, c = 111.1 A). This crystal (Zn-ApUp-Orn) belongs to the P6(5) space group and has three oligoribonuclease molecules per asymmetric unit.
Fiedler
Tristan J
TJ
Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, PO Box 016129, Miami, FL 33101-6129, USA.
Vincent
Helen A
HA
Zuo
Yuhong
Y
Gavrialov
Orit
O
Malhotra
Arun
A
D003001:Cloning, Molecular; D003460:Crystallization / Q000379:methods; D018360:Crystallography, X-Ray; D019281:Dimerization; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics; D005095:Exoribonucleases / Q000737:chemistry* / Q000235:genetics; D008345:Manganese
D029968:Escherichia coli Proteins; D008345:Manganese; D005095:Exoribonucleases; C094587:oligoribonuclease