title: Recombinant expression and purification of functional XorII, a restriction endonuclease from Xanthomonas oryzae pv. oryzae.
abstract: An endonuclease from Xanthomonas oryzae pathovar oryzae KACC 10331, XorII, was recombinantly produced in Escherichia coli using a T7 system. XorII was purified using a combination of ion exchange and immobilized metal affinity chromatography (IMAC). An optimized washing protocol was carried out on an IMAC in order to obtain a high purity product. The final amount of purified XorII was approximately 2.5 mg/L of LB medium. The purified recombinant XorII was functional and showed the same cleavage pattern as PvuI. The enzyme activity tested the highest at 25 degrees in 50 mM NaCl, 10 mM Tris-HCl, 10 mM MgCl2, and 1 mM dithiothreitol at a pH of 7.9.
authors.affiliation: Department of Chemistry, Sejong University, Seoul 143-747, Republic of Korea.
authors.forename: Dong Kyu
authors.initials: DK
authors.lastname: Hwang
authors.forename: Jae-Yong
authors.initials: JY
authors.lastname: Cho
authors.forename: Young Kee
authors.initials: YK
authors.lastname: Chae
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D004262:DNA Restriction Enzymes / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D014974:Xanthomonas / Q000201:enzymology* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D004262:DNA Restriction Enzymes