FIG. 5.

Complex formation of Orf22 and GAM-1 with pRb mutants. (A) CEF cells were transfected with 3 μg of the indicated expression constructs and lysed 24 h after transfection. Equal amounts of the expressed proteins were incubated with recombinant GST-Rb proteins as described in Materials and Methods. Complex formation was monitored by subsequent Western analysis with anti-Myc antibody. (B) Competition assay with T peptide. Myc-tagged constructs of Orf22, GAM-1, and E1A 19K were expressed in CEF cells. Lysates of these cells were incubated with recombinant GST-Rb together with increasing amounts of T peptide. Lane 1, no peptide; lane 2, 20 μg of peptide; lane 3, 40 μg of peptide; lane 4, lysate of transfected CEF cells (40 μl).