cdc2 specifically binds to the carboxyl terminus of FAC. (A) GST fusion proteins used for in vitro mixing experiments are shown schematically. (B) Whole cell extract (unlabeled) was made from HeLa cells first synchronized at G₁ /S by double thymidine block and then released for 8 hours in nocodazole.41 GST (lane 1), GST-N (lane 2), GST-FAC (wild-type; lane 3), GST-FAC (L554P, lane 4), or GST-C1 (lane 5), prebound to glutathione-sepharose beads, was mixed with the extract. Bound cellular proteins were electrophoresed by SDS-PAGE and transferred to nitrocellulose. The filter was probed with either anti-GST monoclonal antibody or anti-cdc2 monoclonal antibody.