Fig. 5. Mutation of serine 345 to alanine or aspartic acid abolishes association of Chk1 with 14-3-3 upon DNA damage. NW223 (Chk1), NW444 (S345A) and NW463 (S345D) were grown to mid-log phase, treated with CPT and lysed. Lysates were incubated with antibody against Rad 24 (UMDNJ 55), subjected to SDS-PAGE and immunoblotted with 12CA5 anti-HA antibody (IP). Control precipitations (NI) with non-immune rabbit serum were processed exactly as anti-Rad24 IPs. Samples of the lysates, before immunoprecipitation are shown in the left panel.