Deciphering the function of lactococcal phage ul36 Sak domains

J Struct Biol. 2010 Jun;170(3):462-9. doi: 10.1016/j.jsb.2009.12.021. Epub 2009 Dec 29.

Abstract

Virulent phages are responsible for milk fermentation failures in the dairy industry, due to their ability to infect starter cultures containing strains of Lactococcus lactis. Single-strand annealing proteins (SSAPs) have been found in several lactococcal phages, among which Sak in the phage ul36. Sak has been recently shown to be a functional homolog of the human protein RAD52, involved in homologous recombination. A comparison between full-length Sak and its N- and C-terminal domains was carried out to elucidate functional characteristics of each domain. We performed HPLC-SEC, AFM and SPR experiments to evaluate oligomerization states and compare the affinities to DNA. We have shown that the N-terminal domain (1-171) is essential and sufficient for oligomerization and binding to DNA, while the C-terminal domain (172-252) does not bind DNA nor oligomerize. Modelisation of Sak N-terminal domain suggests that DNA may bind a positively charged crevice that runs external to the ring. Annealing and stimulation of RecA strand exchange indicate that only the N-terminal domain is capable of single-strand annealing and both domains do not stimulate the RecA strand exchange reaction. We propose that Sak N-terminus is involved in DNA binding and annealing while the C-terminus may serve to contact Sak partners.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / physiology
  • Bacteriophages / chemistry*
  • Bacteriophages / genetics
  • Bacteriophages / pathogenicity
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • Humans
  • Lactococcus lactis / physiology
  • Lactococcus lactis / virology*
  • Microscopy, Atomic Force
  • Models, Molecular
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / physiology
  • Protein Structure, Tertiary
  • Rec A Recombinases / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Structural Homology, Protein
  • Surface Plasmon Resonance
  • Viral Proteins / chemistry*
  • Viral Proteins / genetics
  • Viral Proteins / physiology

Substances

  • AbiK protein, Lactococcus lactis
  • Bacterial Proteins
  • DNA-Binding Proteins
  • Peptide Fragments
  • RDM1 protein, human
  • Recombinant Proteins
  • Viral Proteins
  • Rec A Recombinases