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Figure 4 | Retrovirology

Figure 4

From: The role of cyclin D2 and p21/waf1 in human T-cell leukemia virus type 1 infected cells

Figure 4

Effect of purified p21/waf1 and p16/INK4A on cyclin D2/cdk4 complex. (A) Recombinant cdk2, cdk4, cyclin E, p21/waf1 wildtype (WT), p21/waf1 mutant in the cyclin binding site (mut), p16/INK4A, and cyclin D2 were expressed and purified using affinity tag chromatography. Following purification an aliquot was separated on by SDS-PAGE on a 4–20% gel and silver stained for purity. Dots (.) represent authentic cell cycle proteins (B) In vitro kinase assays with purified cyclin D2, cyclin E, cdk4, cdk2, p16/INK4A, p21/waf1 (WT) and p21/waf1 (mut) were performed using GST-Rb as a substrate for 1 hour at 37°C and processed as described in the methods section. One hundred nanograms of cdk4, cyclin D2, p16/INK4A, cdk2, and cyclin E were used in the kinase assays. (C) In vitro kinase assays were performed using GST-Rb as described in the methods section. One hundred nanograms of cdk4 and cyclin D2 were used in the kinase assays. (D) In vitro kinase assays were performed using GST-Rb as described above. Concentrations of flavopiridol used were 10, 50, and 100 nM for lanes 2–4, respectively.

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