A structural phylogenetic tree of Rad52 and its annealase superfamily.

Rad52, a highly conserved eukaryotic protein, plays a crucial role in DNA repair, particularly in double-strand break repair. Recent findings reveal that its distinct structural features, including a characteristic β-sheet and β-hairpin motif, are shared with the lambda phage single-strand annealing protein, Redβ, and other prokaryotic single-strand annealing proteins (SSAPs), indicating a common superfamily. Our analysis of over 10,000 SSAPs across all domains of life supports this hypothesis, confirming the presence of the characteristic motif despite variations in size and composition. We found that archaea, representing only 1% of the studied proteins, exhibit most of these variations as reflected by their spread across the phylogenetic tree, whereas eukaryotes exhibit only Rad52. By examining four representative archaeal SSAPs, we elucidate the structural relationship between eukaryotic and bacterial SSAPs, highlighting differences in β-sheet size and β-hairpin complexity. Furthermore, we identify an archaeal SSAP with a predicted structure nearly identical to human Rad52. Together with a screen of over 100 million unannotated proteins for potential SSAP candidates, our computational analysis complements the existing sequence and structural evidence supporting orthology among five SSAP families: Rad52, Redβ, RecT, Erf, and Sak3.