Fig. 7

In vitro autophosphorylation of wild-type and mutant GlPLKs. a A schematic diagram of GlPLK. Serine/threonine kinase (KD) and two polo-box domains (PBD1, PBD2) are indicated as boxes. Lys51 (K51) is suggested as a residue that initially receives phosphate from ATP, and two threonine residues (T179 and T183) are proposed as the target sites of subsequent phosphorylation. b Western blot of rGlPLK proteins synthesized in vitro using mouse anti-c-Myc antibodies (1:1000). c Phosphorylation of in vitro-synthesized GlPLKs. The c-Myc-tagged GlPLKs (wild-type GlPLK, K51R mutant GlPLK, and T179AT183A mutant GlPLK) were prepared in vitro and then used for kinase assays. GlPLK proteins were resuspended in 20 μl kinase buffer in the presence of 2.5 µCi [γ-³²P]ATP. The kinase reactions were then subjected to 12% SDS-PAGE and visualized by autoradiography