CDK1/cyclin B phosphorylates Ubc9 <i>in vitro</i>.

Abstract

<p>(A) <i>In vitro</i> phosphorylation of SAE1/SAE2 by CDK1/cyclin B. CDK1/cyclin B (3 nM, 6 nM and 12 nM) was incubated with (left) or without (right) SAE1/His₆-SAE2 for 30 min at 30°C in the presence of [γ-³²P] ATP. The reactions were analyzed by 10% SDS-PAGE followed by Coomassie blue staining and autoradiography. (B) <i>In vitro</i> phosphorylation of His₆-SUMO1 by CDK1/cyclin B. Various concentrations of CDK1/cyclin B (as mentioned above) were incubated with (left) or without His₆-SUMO1 (right) in the presence of [γ-³²P] ATP for 30 min at 30°C. (C) <i>In vitro</i> phosphorylation of Ubc9 by CDK1/cyclin B. Various concentrations of CDK1/cyclin B (as mentioned above) were incubated with (left panel) or without His₆-Ubc9 (right panel) in the presence of [γ-³²P] ATP for 30 min at 30°C. (D) <i>In vitro</i> phosphorylation of GST-hTOP1 ^(110–125)AA. Various concentrations of CDK1/cyclin B (as mentioned above) were incubated with (left) or without GST-hTOP1^(110–125)AA (right) in the presence of [γ-³²P] ATP. Reaction mixtures of B, C and D were analyzed by 15% SDS-PAGE followed by Coomassie blue staining and autoradiography.</p