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. Author manuscript; available in PMC: 2009 Aug 18.
Published in final edited form as: Cancer Res. 2007 Jul 15;67(14):6685–6690. doi: 10.1158/0008-5472.CAN-07-0495

Fig. 4.

Fig. 4

CDK2-cyclinA phosphorylates ATRIP S224. (A) Recombinant His-MBP-ATRIP or His-MBP-ATRIP S224A were purified from bacterial cells and added to an in vitro kinase assay with recombinant CDK2-cyclin A. Following incubation, the kinase reaction was separated on SDS-PAGE and stained. The autoradiogram showing the amount of 32P incorporated into the ATRIP proteins and a coomassie stain of the gel demonstrating that equal amounts of substrate were added to each reaction is shown. A reaction in which no substrate was added is included as a control (−). (B) Wild-type or mutant HA-ATRIP-Flag-ATR complexes were purified from transiently transfected HEK293T cells with HA-agarose beads. Kinase reactions were performed with recombinant CDK2-cyclin A complexes. Reactions were separated by SDS-PAGE. Equivalent gels were stained with coomassie blue and exposed to film or immunoblotted with antibodies to ATRIP and ATR to visualize the amount of the ATR-ATRIP complex in each reaction.