DISSECTION OF THE METHYL-CPG BINDING DOMAIN FROM THE CHROMOSOMAL PROTEIN MECP2

MeCP2 is a chromosomal protein which binds to DNA that is methylated a t CpG. In situ immunofluorescence in mouse cells has shown that the pr otein is most concentrated in pericentromeric heterochromatin, suggest ing that MeCP2 may play a role in the formation of inert chromatin. He re we have isolated a minimal methyl-CpG binding domain (MBD) from MeC P2. MBD is 85 amino acids in length, and binds exclusively to DNA that contains one or more symmetrically methylated CpGs. MBD has negligabl e non-specific affinity for DNA, confirming that non-specific and meth yl-CpG specific binding domains of MeCP2 are distinct. In vitro footpr inting indicates that MBD binding can protect a 12 nucleotide region s urrounding a methyl-CpG pair, with an approximate dissociation constan t of 10(-9)M.