title: Rett syndrome-causing mutations in human MeCP2 result in diverse structural changes that impact folding and DNA interactions.
abstract: Most cases of Rett syndrome (RTT) are caused by mutations in the methylated DNA-binding protein, MeCP2. Here, we have shown that frequent RTT-causing missense mutations (R106W, R133C, F155S, T158M) located in the methylated DNA-binding domain (MBD) of MeCP2 have profound and diverse effects on its structure, stability, and DNA-binding properties. Fluorescence spectroscopy, which reports on the single tryptophan in the MBD, indicated that this residue is strongly protected from the aqueous environment in the wild type but is more exposed in the R133C and F155S mutations. In the mutant proteins R133C, F155S, and T158M, the thermal stability of the domain was strongly reduced. Thermal stability of the wild-type protein was increased in the presence of unmethylated DNA and was further enhanced by DNA methylation. DNA-induced thermal stability was also seen, but to a lesser extent, in each of the mutant proteins. Circular dichroism (CD) of the MBD revealed differences in the secondary structure of the four mutants. Upon binding to methylated DNA, the wild type showed a subtle but reproducible increase in alpha-helical structure, whereas the F155S and R106W did not acquire secondary structure with DNA. Each of the mutant proteins studied is unique in terms of the properties of the MBD and the structural changes induced by DNA binding. For each mutation, we examined the extent to which the magnitude of these differences correlated with the severity of RTT patient symptoms.
authors.affiliation: Department of Biology, Program in Molecular and Cellular Biology, University of Massachusetts, Amherst, MA 01003, USA.
authors.forename: Rajarshi P
authors.initials: RP
authors.lastname: Ghosh
authors.forename: Rachel A
authors.initials: RA
authors.lastname: Horowitz-Scherer
authors.forename: Tatiana
authors.initials: T
authors.lastname: Nikitina
authors.forename: Lila M
authors.initials: LM
authors.lastname: Gierasch
authors.forename: Christopher L
authors.initials: CL
authors.lastname: Woodcock
mesh_terms: D002942:Circular Dichroism; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D009691:Nucleic Acid Denaturation; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D015518:Rett Syndrome / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
chemical_list: D051783:Methyl-CpG-Binding Protein 2; D004247:DNA