The sequence specificity domain of cytosine-C5 methylases

Nucleic Acids Res. 1991 Nov 25;19(22):6183-90. doi: 10.1093/nar/19.22.6183.


Prokaryotic DNA[cytosine-C5]methyltransferases (m5C-methylases) share a common architectural arrangement of ten conserved sequence motifs. A series of eleven hybrids have been constructed between the HpaII (recognition sequence: Cm5CGG) and HhaI (recognition sequence: Gm5CGC) DNA-methylases. The hybrids were over-expressed in E.coli and their in vivo methylation phenotypes investigated. Six were inactive by our assay while five of them retained partial methylation activity and full specificity. In all five cases the specificity matched that of the parent methylase which contributed the so-called variable region, located between conserved motifs VIII and IX. This was the only sequence held in common between the active hybrids and for the first time provides unequivocal evidence that the specificity determinants of the mono-specific m5C-methylases are located within the variable region. Correlation of the hybrid methylase structure with the efficiency of methylation suggests that conserved motif IX may interact with the variable region whereas motif X most probably interacts with the N-terminal half of the molecule.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • DNA Restriction Enzymes
  • DNA, Bacterial / genetics
  • DNA-Cytosine Methylases / genetics*
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Genes, Bacterial
  • Methylation
  • Molecular Sequence Data
  • Mutagenesis
  • Plasmids
  • Polymerase Chain Reaction
  • Sequence Alignment


  • DNA, Bacterial
  • DNA-Cytosine Methylases
  • DNA Restriction Enzymes