Poster Presentation The 47th Lorne Conference on Protein Structure and Function 2022

Half a century after its discovery, the cryo-EM structure of Erf annealase of phage P22 provides insights into its evolution and molecular mechanisms (#153)

MariaKatarina Lambourne 1 2 , Jodi Brewster 1 2 , Gökhan Tolun 1 2
  1. School of Chemistry and Molecular Biosciences and Molecular Horizons Institute, University of Wollongong, Wollongong, NSW, Australia
  2. Illawarra Health and Medical Research Institute, Wollongong, NSW, Australia

Single-strand annealing homologous DNA recombination (SSA) is a process conserved throughout evolution from bacteriophages to humans1, highlighting its importance and indispensability. It is particularly vital in viruses, as it catalyses circularisation and concatamerisation of the viral DNA, in addition to repairing the damaged viral genomes2.

SSA is catalysed by EATR (Exonuclease Annealase Two-component Recombinase) complexes. Exonuclease catalyses digestion of one of the DNA strands in the 5ʹ to 3ʹ direction, generating a 3′ ssDNA overhang. The annealase binds to the ssDNA and catalyses homology searching and annealing between homologous DNAs. Despite half a century of extensive research into SSA and EATRs, the molecular mechanistic details are poorly understood3.

Bacteriophage P22 is a dsDNA virus that infects Salmonella enterica s. Typhimurium, and encodes its own recombination system. Erf (essential recombination function) protein is the annealase of phage P22, which is the defining member of the Erf protein family. Despite its discovery in 19704, a structure of Erf was still not available.

We report the first atomic structure of Erf, more than 50 years after its discovery. The cryo-EM map reconstructed at 2.47 Å resolution shows Erf forming a nonadecameric ring structure (a homo-oligomer composed of 19 subunits). The atomic model of Erf reveals a striking similarity to the human Rad52 annealase, containing a β-β-β-α fold, which most likely forms the DNA-binding site. Upon homo-oligomerisation, this site forms a positively charged groove that can accommodate ssDNA binding. We discuss the implications of our findings from both evolutionary and molecular mechanistic perspectives.

  1. San Filippo, J., Sung, P. & Klein, H. Mechanism of eukaryotic homologous recombination. Annual Review of Biochemistry. vol. 77 (2008).
  2. Krejci, L., Altmannova, V., Spirek, M. & Zhao, X. Homologous recombination and its regulation. Nucleic Acids Research. vol. 40, 5795–5818 (2012).
  3. Brewster, J. L. & Tolun, G. Half a century of bacteriophage lambda recombinase: In vitro studies of lambda exonuclease and Red-beta annealase. IUBMB Life. vol. 72, 1622–1633 (2020).
  4. Botstein, D. & Matz, M. J. A recombination function essential to the growth of bacteriophage P22. Journal of Molecular Biology. vol. 54, 417–440 (1970).