Cloning and characterization of the unusual restriction-modification system comprising two restriction endonucleases...

An Escherichia coli RFL47 DNA fragment containing the Eco47IR and Eco47II restriction-modification (R-M) system has been cloned and sequenced. A clone carrying this system has been selected by its ability to restrict phage lambda in vivo. The sequence of 5360 bp was determined, and its analysis revealed three major open reading frames (ORF) corresponding to two restriction endonucleases (ENases) and one DNA methyltransferase (MTase): R.Eco47II (239 amino acid (aa)), R.Eco47I (230 aa) and M.Eco47II (417 aa). The M.Eco47II aa sequence possesses all conserved domains typical for m5C MTases and its variable region has a high homology with M.Sau96I and M.SinI. The ORF harboring a predicted helix-turn-helix motif upstream from the eco47IR gene has been found. No sequence resembling the eco47IM gene has been detected in the complete fragment sequenced, although disrupted ORF, possibly corresponding to the transposase-encoding gene, has been found in the intergenic area between eco47IIM and eco47IR. No homology was found between the ENases; however, both revealed homology with their isoschizomers, R.SinI and R.Sau96I.