DEV024919 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. The Cdk1^12aa-Cdk2^Δ12 fusion protein is as active as wild-type Cdk2. The fusion protein encoded by the first coding exon of the Cdk1 locus (amino acids 1-12) and the Cdk2^Δ12-HA cDNA contains four conservative changes as compared with wild-type Cdk2: N3D, F4Y, Q5I and V7I (see also Fig. 1A). We generated an HA-tagged Cdk2 construct containing these mutations and transfected it along with wild-type Cdk2-HA and Myc-tagged cyclin E1 or E2 into 293 cells. After preparing protein extracts, Myc-cyclin E was immunoprecipitated (in a complex with wild-type or mutant Cdk2) and assayed for in vitro kinase activity using histone H1 as a substrate in the presence of radiolabeled ATP. This experiment indicates that Cdk2^N3DF4YQ5IV7I-HA displays similar levels of activity to wild-type Cdk2.