Molecular characterization of a phage-encoded resistance system in Lactococcus lactis

A specific fragment of the genome of Tuc2009, a temperate lactococcal bacte riophage, was shown to contain several open reading frames, whose deduced p rotein products exhibited similarities to proteins known to be involved in DNA replication and modification. In this way, a putative single-stranded b inding protein, replisome organizer protein, topoisomerase I, and a methyla se were identified. When the genetic information coding for the putative re plisome organizer protein of Tuc2009, Rep(2009), was supplied on a high-cop y-number plasmid vector, it was shown to confer a phage-encoded resistance (Per) phenotype on its lactococcal host UC509.9, The presence of this recom binant plasmid was shown to cause a marked reduction in Tuc2009 DNA replica tion, suggesting that the observed phage resistance was due to titration of a factor, or factors, required for Tuc2009 DNA replication. Further experi ments delineated the phage resistance-conferring region to a 160-bp fragmen t rich in direct repeats. Gel retardation experiments, which indicated a pr otein-DNA interaction between this 160-bp fragment and the Rep(2009) protei n, were performed. UC509.9 strains harboring plasmids with randomly mutated versions of this fragment were shown to display a variable phage resistanc e phenotype, depending on the position of the mutations.