Conformational adaptability of Redbeta during DNA annealing and implications for its structural relationship with Rad52

• Published in: Journal of molecular biology Vol. 391; no. 3; pp. 586 - 598
• Main Authors: Erler, Axel, Wegmann, Susanne, Elie-Caille, Celine, Bradshaw, Charles Richard, Maresca, Marcello, Seidel, Ralf, Habermann, Bianca, Muller, Daniel J, Stewart, A Francis
• Format: Journal Article
• Language: English
• Published: England 21.08.2009
• Subjects: Base Pairing; Binding Sites; DNA, Single-Stranded - chemistry; DNA, Single-Stranded - ultrastructure; Microscopy, Atomic Force; Protein Structure, Quaternary; Rad52 DNA Repair and Recombination Protein - chemistry; Rad52 DNA Repair and Recombination Protein - metabolism; Rad52 DNA Repair and Recombination Protein - ultrastructure; Recombination, Genetic
• ISSN: 1089-8638
• DOI: 10.1016/j.jmb.2009.06.030

Single-strand annealing proteins, such as Redbeta from lambda phage or eukaryotic Rad52, play roles in homologous recombination. Here, we use atomic force microscopy to examine Redbeta quaternary structure and Redbeta-DNA complexes. In the absence of DNA, Redbeta forms a shallow right-handed helix. The presence of single-stranded DNA (ssDNA) disrupts this structure. Upon addition of a second complementary ssDNA, annealing generates a left-handed helix that incorporates 14 Redbeta monomers per helical turn, with each Redbeta monomer annealing approximately 11 bp of DNA. The smallest stable annealing intermediate requires 20 bp DNA and two Redbeta monomers. Hence, we propose that Redbeta promotes base pairing by first increasing the number of transient interactions between ssDNAs. Then, annealing is promoted by the binding of a second Redbeta monomer, which nucleates the formation of a stable annealing intermediate. Using threading, we identify sequence similarities between the RecT/Redbeta and the Rad52 families, which strengthens previous suggestions, based on similarities of their quaternary structures, that they share a common mode of action. Hence, our findings have implications for a common mechanism of DNA annealing mediated by single-strand annealing proteins including Rad52.