Solution Structure of the Methyl-CpG Binding Domain of Human MBD1 in Complex with Methylated DNA

Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, 630-0101, Nara, Japan.
Cell (Impact Factor: 32.24). 06/2001; 105(4):487-97. DOI: 10.1016/S0092-8674(01)00324-5
Source: PubMed


In vertebrates, the biological consequences of DNA methylation are often mediated by protein factors containing conserved methyl-CpG binding domains (MBDs). Mutations in the MBD protein MeCP2 cause the neurodevelopmental disease Rett syndrome. We report here the solution structure of the MBD of the human methylation-dependent transcriptional regulator MBD1 bound to methylated DNA. DNA binding causes a loop in MBD1 to fold into a major and novel DNA binding interface. Recognition of the methyl groups and CG sequence at the methylation site is due to five highly conserved residues that form a hydrophobic patch. The structure indicates how MBD may access nucleosomal DNA without encountering steric interference from core histones, and provides a basis to interpret mutations linked to Rett syndrome in MeCP2.

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Available from: Izuru Ohki, Feb 10, 2015
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    • "All of the genes in this region are of special interest because of their known functions and/or their relationship (association) with neuropsychiatric disorders. MBD1 is known to be associated with Rett syndrome and autism spectrum disorders.54,55) The ME2 gene is shown to be associated with idiopathic generalized epilepsy; the strongest association was seen with variants in the promoter region area of this gene.56) "
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    ABSTRACT: Objective It was previously suggested that the malic enzyme 2 (ME2) as the candidate gene for psychosis in fine mapping of chromosome 18q21. Chromosome 18q21 is also one of the possible regions that can contribute to addiction. Methods We performed a pilot study for discovering candidate gene of chromosome 18q21 in the methamphetamine abusers for elucidating the candidate gene for methamphetamine addiction leading to psychosis. We have selected 30 unrelated controls (16 males, 14 females; age=59.8±10.4) and 37 male methamphetamine abusers (age=43.3±7.8). We analyzed 20 single nucleotide polymorphisms (SNPs) of 7 neuronal genes in chromosome 18q21 for DNA samples that was checked for the data quality and genotype error. The association between the case-control status and each individual SNP was measured using multiple logistic regression models (adjusting for age and sex as covariates). And we controlled false discovery rate (FDR) to deal with multiple testing problem. Results We found 3 significant SNPs of 2 genes in chromosome 18q21 (p-value<0.05; adjusting for age as covariate) in methamphetamine abusers compared to controls. We also found 2 significant SNPs of 1 gene (p-value<0.05; adjusting for age and sex as covariates) (rs3794899, rs3794901:MAPK4). Two SNPs in MAPK4 gene were significant in both statistical groups. Conclusion MAPK4, the gene for mitogen-activated protein kinase 4, is one of the final 6 candidate genes including ME2 in 18q12-21 in our previous finemapping for psychosis. Our results suggest that MAPK4 can be a candidate gene that contribute to the methamphetamine addiction leading to psychosis.
    Clinical Psychopharmacology and Neuroscience 04/2014; 12(1):54-64. DOI:10.9758/cpn.2014.12.1.54
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    • "Based on these studies, the methyl binding proteins/enzymes can be divided into two broad groups, depending on whether they recognize the methyl group in the context of double stranded DNA or whether they flip the modified base to scrutinize it in a dedicated pocket. MBDs (1–4), MeCP2 (4) and Kaiso (5) interact with the modified base in a Watson-Crick pair. In contrast, UHRF1 and most likely also MspJI share a so-called SRA (SET and RING associated) domain that flips and accommodates the modified base (6–8,10). "
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    ABSTRACT: PvuRts1I is a prototype for a larger family of restriction endonucleases that cleave DNA containing 5-hydroxymethylcytosine (5hmC) or 5-glucosylhydroxymethylcytosine (5ghmC), but not 5-methylcytosine (5mC) or cytosine. Here, we report a crystal structure of the enzyme at 2.35 Å resolution. Although the protein has been crystallized in the absence of DNA, the structure is very informative. It shows that PvuRts1I consists of an N-terminal, atypical PD-(D/E)XK catalytic domain and a C-terminal SRA domain that might accommodate a flipped 5hmC or 5ghmC base. Changes to predicted catalytic residues of the PD-(D/E)XK domain or to the putative pocket for a flipped base abolish catalytic activity. Surprisingly, fluorescence changes indicative of base flipping are not observed when PvuRts1I is added to DNA substrates containing pyrrolocytosine in place of 5hmC (5ghmC). Despite this caveat, the structure suggests a model for PvuRts1I activity and presents opportunities for protein engineering to alter the enzyme properties for biotechnological applications.
    Nucleic Acids Research 03/2014; 42(9). DOI:10.1093/nar/gku186 · 9.11 Impact Factor
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    • "Precipitation of the DNA fragments containing 5mC was performed as described previously [32] with a slight modification. In brief, 10 μg of sonicated DNA was incubated with 1.2 μg of recombinant His-GST-MBD1 coding 1-75 of MBD1 [33] and MagneGST beads (Promega) at 4°C overnight. Bound DNA was eluted by proteinase K treatment at 50°C for 3 h. "
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    ABSTRACT: Hydroxymethylcytosine in the genome is reported to be an intermediate of demethylation. In the present study, we demonstrated that maintenance methyltransferase Dnmt1 scarcely catalyzed hemi-hydroxymethylated DNA and that the hemi-hydroxymethylated DNA was not selectively recognized by the SRA domain of Uhrf1, indicating that hydroxymethylcytosine is diluted in a replication-dependent manner. A high level of 5-hydroxymethylcytosine in mouse embryonic stem cells was produced from the methylcytosine supplied mainly by de novo-type DNA methyltransferases Dnmt3a and Dnmt3b. The promoter regions of the HoxA gene cluster showed a high hydroxymethylation level whilst the methylcytosine level was quite low, suggesting that methylated CpG is actively hydroxylated during proliferation. All the results indicate that removal and production of hydroxymethylcytosine are regulated in replication-dependent manners in mouse embryonic stem cells.
    PLoS ONE 12/2013; 8(12):e82961. DOI:10.1371/journal.pone.0082961 · 3.23 Impact Factor
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