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.

Deciphering the function of lactococcal phage ul36 Sak domains / Scaltriti, Erika; S., Moineau; H., Launay; J. Y., Masson; Rivetti, Claudio; Ramoni, Roberto; V., Campanacci; Tegoni, Mariella; C., Cambillau. - In: JOURNAL OF STRUCTURAL BIOLOGY. - ISSN 1047-8477. - 170(3):(2010), pp. 462-469. [10.1016/j.jsb.2009.12.021]

Deciphering the function of lactococcal phage ul36 Sak domains.

SCALTRITI, Erika;RIVETTI, Claudio;RAMONI, Roberto;TEGONI, Mariella;
2010

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.
Deciphering the function of lactococcal phage ul36 Sak domains / Scaltriti, Erika; S., Moineau; H., Launay; J. Y., Masson; Rivetti, Claudio; Ramoni, Roberto; V., Campanacci; Tegoni, Mariella; C., Cambillau. - In: JOURNAL OF STRUCTURAL BIOLOGY. - ISSN 1047-8477. - 170(3):(2010), pp. 462-469. [10.1016/j.jsb.2009.12.021]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11381/2307588
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