Figure 3: EmMBD2 selectively binds methylated DNA.
From: Methylation specific targeting of a chromatin remodeling complex from sponges to humans
(a) Surface plasmon resonance analyses shows that EmMBD2/3 binds with similar affinity and selectivity for methylated (mCpG) and unmethylated (CpG(x3)) DNA as GgMBD2. (b) 2D 15N-HSQC spectra show similar chemical shift distributions for EmMBD2/3 and GgMBD2 with very distinct resonances for reporter residues G35 and A38 (circled and labeled). (c) As we described previously14, the unusual chemical shifts for the sidechain of R32(15Nε1Hε) and the backbone amides of G35 and A38 show large differences between mCpG specific (red) and non-specific (cyan) binding modes. These three residues, shown as red spheres in the cartoon diagram of EmMBD2/3 MBD, are on a loop connecting the central β-strands of the MBD. R32 forms critical bidentate hydrogen bonds with a guanosine base in the CpG dinucleotide and with the sidechain of D42 (see Fig. 2) which stabilizes this loop. (d) 2D 15N-HSQC spectra show that resonances for these three residues in EmMBD2/3 show large, and proportional chemical shift changes whether bound to methylated (mCpG), unmethylated (CpG(x3)), or DNA that lacks any CpG dinucleotides (no CpG). This behavior is very similar to what we previously described for GgMBD214 and indicates that EmMBD2/3 preferentially localizes to densely methylated regions.
