title: DNA binding restricts the intrinsic conformational flexibility of methyl CpG binding protein 2 (MeCP2).
abstract: Mass spectrometry-based hydrogen/deuterium exchange (H/DX) has been used to define the polypeptide backbone dynamics of full-length methyl CpG binding protein 2 (MeCP2) when free in solution and when bound to unmethylated and methylated DNA. Essentially the entire MeCP2 polypeptide chain underwent H/DX at rates faster than could be measured (i.e. complete exchange in ≤10 s), with the exception of the methyl DNA binding domain (MBD). Even the H/DX of the MBD was rapid compared with that of a typical globular protein. Thus, there is no single tertiary structure of MeCP2. Rather, the full-length protein rapidly samples many different conformations when free in solution. When MeCP2 binds to unmethylated DNA, H/DX is slowed several orders of magnitude throughout the MBD. Binding of MeCP2 to methylated DNA led to additional minor H/DX protection, and only locally within the N-terminal portion of the MBD. H/DX also was used to examine the structural dynamics of the isolated MBD carrying three frequent mutations associated with Rett syndrome. The effects of the mutations ranged from very little (R106W) to a substantial increase in conformational sampling (F155S). Our H/DX results have yielded fine resolution mapping of the structure of full-length MeCP2 in the absence and presence of DNA, provided a biochemical basis for understanding MeCP2 function in normal cells, and predicted potential approaches for the treatment of a subset of RTT cases caused by point mutations that destabilize the MBD.
authors.affiliation: Department of Biochemistry and Molecular Biology, Colorado State University, Fort Collins, Colorado 80523-1870, USA. jeffrey.c.hansen@colostate.edu
authors.forename: Jeffrey C
authors.initials: JC
authors.lastname: Hansen
authors.forename: Brian B
authors.initials: BB
authors.lastname: Wexler
authors.forename: Danielle J
authors.initials: DJ
authors.lastname: Rogers
authors.forename: Kristopher C
authors.initials: KC
authors.lastname: Hite
authors.forename: Tanya
authors.initials: T
authors.lastname: Panchenko
authors.forename: Sandya
authors.initials: S
authors.lastname: Ajith
authors.forename: Ben E
authors.initials: BE
authors.lastname: Black
mesh_terms: D019943:Amino Acid Substitution; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D010449:Peptide Mapping; D011485:Protein Binding; D011487:Protein Conformation; D055550:Protein Stability; D015518:Rett Syndrome / Q000235:genetics / Q000378:metabolism
chemical_list: C497392:MECP2 protein, human; D051783:Methyl-CpG-Binding Protein 2; D004247:DNA