The NgoPII restriction endonuclease, which recognizes the sequence 5-GGCC-3, differs from its isoschizomer HaeIII in being sensitive to methylation at the external cytosine residue. The entire nucleotide sequence of a cloned 3.3 kb segment of Neisseria gonorrhoeae strain P9 chromosomal DNA which harbours the NgoPII restriction-modification system has been determined. This data, coupled with sub-cloning experiments, indicates that the restriction endonuclease (R.NgoII) and modification (M.NgoII) genes are transcribed from separate promoters but are arranged in tandem, with the R.NgoPII gene being located on the 5 side of the M.NgoPII gene. Unlike all previously reported restriction systems the 3 end of the endonuclease open reading frame overlaps the 5 end of the methylase open reading frame by 8 codons. This overlap may have implications for the regulation of the NgoPII restriction-modification system.