We have compared the deduced amino acid (aa) sequences of the EcoRII restriction endonuclease (R-EcoRII) and the proposed specificity (target recognition) domains of three DNA-[cytosine-C5] methyltransferases (MTases), M.EcoRII, M.Dcm, and M.SPR, each of which recognizes the same nucleotide sequence, CCWGG (where W is A or T). We have identified a region containing sequence motifs that are partially conserved in the MTases and R.EcoRII. This may be the first example of aa sequence homology between a MTase specificity (target recognition) domain and its cognate restriction endonuclease (ENase). It suggests that this region is important for DNA recognition by R.EcoRII and that the EcoRII ENase and MTase genes may have evolved from a common progenitor.