Solution structure and intramolecular exchange of methyl-cytosine binding domain protein 4 (MBD4) on DNA suggests a mechanism to scan for mCpG/TpG mismatches.

Unlike other members of the methyl-cytosine binding domain (MBD) family, MBD4 serves as a potent DNA glycosylase in DNA mismatch repair specifically targeting (m)CpG/TpG mismatches arising from spontaneous deamination of methyl-cytosine. The protein contains an N-terminal MBD (MBD4(MBD)) and a C-terminal glycosylase domain (MBD4(GD)) separated by a long linker. This arrangement suggests that the MBD4(MBD) either directly augments enzymatic catalysis by the MBD4(GD) or targets the protein to regions enriched for (m)CpG/TpG mismatches. Here we present structural and dynamic studies of MBD4(MBD) bound to dsDNA. We show that MBD4(MBD) binds with a modest preference for (m)CpG as compared to mismatch, unmethylated and hydroxymethylated DNA. We find that while MBD4(MBD) exhibits slow exchange between molecules of DNA (intermolecular exchange), the domain exhibits fast exchange between two sites in the same molecule of dsDNA (intramolecular exchange). Introducing a single-strand defect between binding sites does not greatly reduce the intramolecular exchange rate, consistent with a local hopping mechanism for moving along the DNA. These results support a model in which the MBD4(MBD4) targets the intact protein to (m)CpG islands and promotes scanning by rapidly exchanging between successive (m)CpG sites which facilitates repair of nearby (m)CpG/TpG mismatches by the glycosylase domain.