Cloning and characterization of filamentous temperature-sensitive protein Z from Xanthomonas oryzae pv. Oryzae

Abstract

The ftsZ gene from Xanthomonas oryzae pv. Oryzae was amplified by PCR with the specific primers, and the recombinant plasmid pET-22b-ftsZ was constructed successfully. The FtsZ with a 6× His tag was overexpressed in a soluble form in Escherichia coli BL21 and purified through a Ni-NTA agarose column. The purified recombinant FtsZ showed a single band on SDS-PAGE with an apparent molecular mass of about 44 kDa, and confirmed by western blotting analysis. The optimum temperature for GTPase activity of the recombined FtsZ was 50 °C, and the optimum pH was 7.0. The recombinant FtsZ showed good stability and retained >95 % activity at 50 °C for 240 min. The GTPase activity followed Michaelis–Menten kinetics with the KM of 1.750 mM and the Vmax of 0.155 nmol Pi/min/nmol FtsZ respectively.