[Cloning and structure analysis of a restriction and modification system, LlaBIII from Lactococcus lactis subsp....

A 22.4 kb naturally occurring plasmid pJW566, isolated from L. lactis W56, was found to encode an R/M system named LlaBIII. The LlaBIII R/M system was isolated on a chloramphenicol resistant derivative of plasmid pJW566, resulting in a plasmid pJK1. Subcloning analysis showed that the LlaBIII determinant was located on a 5 kb HindIII-Sph I fragment. The fragment was sequenced. It contained a single open reading frame (ORF), corresponding to a protein of 1584 or 1576 aa. In the deduced amino acid sequence seven helicase motifs characteristic of endonuclease type I and type III and a conserved catalysis motif X in the R subunits of type I R/M systems were located in the N-terminus, followed by four conserved motifs found in DNA N6-adenine methyltransferases. The C-terminus of the deduced amino acid sequence showed no homology to known R/M systems. Therefore, this polypeptide encoded by LlaBIII is a multifunctional protein possessing putative DNA recognition, methylation and restriction activities.