Figure 4

Effect of R18 on cruciform-binding activity. Lysate was prepared from yeast cells expressing wild-type 14-3-3ε as described in Methods under the WCE preparation section. A- Total protein extract (1 mg) was incubated at 30°C for 30 min in the absence or presence of 100 μM of R18 peptide, followed by immunoprecipitation with anti 14-3-3ε antibody and blotted with anti-Ras2 antibody. Immunoprecipitation with normal rabbit serum (NRS) and a Bmh1/2-expressing strain were used as controls. B- 14-3-3ε protein extracts were incubated in the same conditions as in section A with increasing amounts (10, 50 and 100 μM) of R18, and then subjected to a cruciform binding assay, as described previously (see Methods). Free cruciform DNA denotes the position of the probe alone.