The assay of GTPase activity for the purified Gtr1 wild-type protein using [α-32P]GTP. (A) – The GTPase activity was assayed as described in Methods. As a control, the assay was also performed in the absence of protein. The nucleotides were separated by thin layer chromatography, and the radioactive spots visualized by phosphorimaging. (B) - Michaelis Menten titration and Lineweaver-Burk plot (insert) of the GTPase activity. Recombinant Gtr1 protein was incubated for 60 min at 37 °C with different concentrations of [α-32P] GTP. The initial rates were plotted versus GTP concentration. Km and Vmax were calculated from the Lineweaver-Burk plot. The correlation coefficient was 0.9933.
Sengottaiyan et al. BMC Biochemistry 2012 13:11 doi:10.1186/1471-2091-13-11