Structural flexibility of SRM1 is essential for yeast Rad51 function in vivo.
The yeast rad51 null mutant was transformed either with the empty control vector pYC2, or with the pYC2-Rad51 expression vectors for wild-type Rad51 or rad51-G143P proteins as indicated. Induction of these proteins was under the control of the RAD51 gene promoter. (A) MMS sensitivity assay was carried out as described before (1). (B) Western blot analysis of the wild-type and G143P mutant proteins. Total cell lysates were separated by SDS-PAGE and then analyzed by Western blotting. Anti-Rad51 antibody (Santa Cruz Biotechnology) and anti-tubulin (Invitrogen) were used for detection of Rad51 and tubulin proteins. Tubulin was used here as a protein loading control. Final detection was performed using the ECL detection system, with the emitted chemiluminescence recorded on X-ray film.