The DNA Binding Preference of RAD52 and RAD59 Proteins IMPLICATIONS FOR RAD52 AND RAD59 PROTEIN FUNCTION IN

We examined the double-stranded DNA (dsDNA) binding preference of the Saccharomyces cerevisiae Rad52 protein and its homologue, the Rad59 protein. In nuclease protection assays both proteins protected an internal sequence and the dsDNA ends equally well. Similarly, using electrophoreticmobility shift assays, we found the affinity of both Rad52 and Rad59 proteins for DNA ends to be comparable with their affinity for internal sequences. The protein-DNA complexes were also directly visu-alized using atomic forcemicroscopy. Both proteins formeddis-crete complexes, whichwere primarily found (90–94%) at inter-nal dsDNA sites. We also measured the DNA end binding behavior of human Rad52 protein and found a slight preference for dsDNAends. Thus, these proteins have no strong preference for dsDNA ends over internal sites, which is inconsistent with their function at a step of dsDNA break repair that precede