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. 2013 May 17;288(27):19715–19725. doi: 10.1074/jbc.M112.449652

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© 2013 by The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 2. — MCM7 can be phosphorylated by cyclin E/Cdk2, not cyclin A/Cdk2, on Ser-121 in vitro. A, in vitro kinase assay. GST or GST MCM7 were incubated with GST-cyclin E and GST-Cdk2 or GST-cyclin A and GST-Cdk2, respectively, in the presence of [γ³²P]ATP. Phosphorylation of MCM7 was assessed by SDS-PAGE followed by autoradiography of the gel. B, potential phosphorylation sites on MCM7 by cyclin/CDKs. C, GST and GST-MCM7 and its mutants were incubated with GST-cyclin E and GST-Cdk2 for an in vitro kinase assay in the presence of [γ³²P]ATP. Coomassie Blue (CBB) gel shows the amount of each protein. The relative intensities of phosphorylated protein were calculated. D, GST-MCM7 (1–129) and GST-MCM7 (1–129) S121A were purified for an in vitro kinase assay with GST-cyclin E and GST-Cdk2.