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Query: EC:2.1.1.37 (DNA methyltransferase)
 4,983 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We overexpressed mouse DNA methyltransferase in murine C2C12 myoblast cells and tested the isolated clones for their ability to differentiate. Significant numbers of the clones showed distinct myotubes 24 h after the isolated transformants had been induced to differentiate, whereas the parent C2C12 cells did not form myotubes at this time point. Transfection of the vacant vector or the plasmid containing the reverse-oriented DNA methyltransferase cDNA did not provide significant numbers of transformants with the accelerated differentiation phenotype, suggesting that the effect is caused by the expression of DNA methyltransferase. The expressions of skeletal muscle myosin and creatine kinase in clones that showed the accelerated differentiation-phenotype were also induced about 24 h earlier and at higher levels relative to the parent C2C12 or the control cells, indicating that the entire process of myogenesis had been accelerated. All the methyltransferase-transfected clones, regardless of their phenotypes, demonstrated about threefold higher DNA methyltransferase activity and higher methylation levels than those of the clones transfected with vector alone or the reverse-oriented plasmid. At the early stage of transfection of the sense-oriented plasmid, high de novo methylation activities were detected. We consider it likely that this high de novo methylation activity is the reason for the high methylation levels and the accelerated myotube formation of the clones transfected with the sense-oriented plasmid. In some transformants which showed the accelerated differentiation phenotype, MyoD1 was already fully expressed under the growth conditions while, in control cells, MyoD1 was expressed at low levels. This elevated level of MyoD1 transcription could account for the accelerated myotube formation observed in the transformants. The methylation state of the HpaII sites in exon 1 through exon 2 of the MyoD1 gene and the expression of the MyoD1 transcript are positively correlated.
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PMID:Overexpression of DNA methyltransferase in myoblast cells accelerates myotube formation. 763 39

Methylation of cytosine residues in DNA plays an important role in regulating gene expression during vertebrate embryonic development. Conversely, disruption of normal patterns of methylation is common in tumors and occurs early in progression of some human cancers. In vertebrates, it appears that the same DNA methyltransferase maintains preexisting patterns of methylation during DNA replication and carries out de novo methylation to create new methylation patterns. There are several indications that inherent signals in DNA structure can act in vivo to initiate or block de novo methylation in adjacent DNA regions. To identify sequences capable of enhancing de novo methylation of DNA in vitro, we designed a series of oligodeoxyribonucleotide substrates with substrate cytosine residues in different sequence contexts. We obtained evidence that some 5-methylcytosine residues in these single-stranded DNAs can stimulate de novo methylation of adjacent sites by murine DNA 5-cytosine methyltransferase as effectively as 5-methylcytosine residues in double-stranded DNA stimulate maintenance methylation. This suggests that double-stranded DNA may not be the primary natural substrate for de novo methylation and that looped single-stranded structures formed during the normal course of DNA replication or repair serve as "nucleation" sites for de novo methylation of adjacent DNA regions.
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PMID:5-Methyl-2'-deoxycytidine in single-stranded DNA can act in cis to signal de novo DNA methylation. 763 94

In prokaryotes, the major role of DNA methylation is to protect host DNA against degradation by restriction enzymes. In eukaryotes, DNA methylation has been implicated in the control of several cellular processes, including differentiation, gene regulation, and embryonic development. Structural work on HhaI DNA methyltransferase demonstrates that the substrate nucleotide is completely flipped out of the helix during the modification reaction and has provided much insight into the enzymatic properties of S-adenosyl-L-methionine (SAM)-dependent DNA-modifying enzymes. Structural comparison of three enzymes, HhaI C5-cytosine methyltransferase, TaqI N6-adenine methyltransferase, and catechol O-methyltransferase, reveals a striking similarity in protein folding and indicates that many SAM-dependent methyltransferases have a common catalytic-domain structure. This feature permits the prediction of tertiary structure for other DNA, RNA, protein, and small-molecule methyltransferases from their amino acid sequences, including the eukaryotic CpG methyltransferases.
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PMID:Structure and function of DNA methyltransferases. 766 18

The (cytosine-5) DNA methyltransferase M.HhaI causes its target cytosine base to be flipped completely out of the DNA helix upon binding. We have investigated the effects of replacing the target cytosine by other, mismatched bases, including adenine, guanine, thymine and uracil. We find that M.HhaI binds more tightly to such mismatched substrates and can even transfer a methyl group to uracil if a G:U mismatch is present. Other mismatched substrates in which the orphan guanine is changed exhibit similar behavior. Overall, the affinity of DNA binding correlates inversely with the stability of the target base pair, while the nature of the target base appears irrelevant for complex formation. The presence of a cofactor analog. S-adenosyl-L-homocysteine, greatly enhances the selectivity of the methyltransferase for cytosine at the target site. We propose that the DNA methyltransferases have evolved from mismatch binding proteins and that base flipping was, and still is, a key element in many DNA-enzyme interactions.
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PMID:M.HhaI binds tightly to substrates containing mismatches at the target base. 775 30

DNA methyltransferase activity, present in low salt extracts of nuclei from young pea shoot apices, has been fractionated into two different species by assaying with model substrates. The CG methyltransferase (an unstable enzyme believed to be of 140 kDa) methylates cytosine only in oligonucleotides with CG and Cl dinucleotide targets while an enzyme of 110 kDa (the CNG methyltransferase) methylates the cytosines in 5'-CAG-3' and 5'-CTG-3' target sequences, especially when hemimethylated, but not in 5'-CCG-3' nor in 5'-CGG-3' target sequences present in oligonucleotides.
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PMID:Distinct CG and CNG DNA methyltransferases in Pisum sativum. 775 18

8-Hydroxyl-2'-deoxyguanosine (also referred to as 8-hydroxyguanine [8-OH-dG] or 7,8-dihydro-8-oxoguanine), a common DNA adduct resulting from injury to DNA via reactive oxygen species, affects the in vitro methylation of nearby cytosine moieties by the human DNA methyltransferase. The exact position of 8-OH-deoxyguanosine relative to a CpG dinucleotide appears important to this effect. Our data indicate that 8-OH-deoxyguanosine diminishes the ability of the methyltransferase to methylate a target cytosine when the 8-OH-deoxyguanosine is one or two nucleotides 3' from the cytosine, on the same strand. On the other hand 8-OH-deoxyguanosine does not diminish the ability of the enzyme to respond to a methyl director (5-methylcytosine) when the 8-OH-deoxyguanosine is on the same strand but one or two nucleotides 3' from the methyl director. Differences in methylation rates as great as 13-fold have been detected using various 8-OH-deoxyguanosine-containing oligonucleotides as substrates in methylation assays. Our findings suggest that oxidative damage of parental strand guanines would permit normal copying of methylation patterns through maintenance methylation, while oxidative damage of guanines in the nascent strand DNA would inhibit such methylation.
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PMID:DNA adduct 8-hydroxyl-2'-deoxyguanosine (8-hydroxyguanine) affects function of human DNA methyltransferase. 776 94

Enzymatic methylation of DNA plays important roles in both prokaryotes and eukaryotes. Structural study of the HhaI DNA methyltransferase has provided considerable insight into the chemistry of C5-cytosine methylation. The DNA-protein complex reveals a substrate cytosine flipped out of the double helix during the reaction, and a novel two-loop DNA-binding motif used for both sequence recognition and flipping the base. Structural comparison of HhaI C5-cytosine methyltransferase, TaqI N6-adenine methyltransferase, and catechol O-methyltransferase reveals a common catalytic domain structure, which might be universal among S-adenosyl-L-methionine (SAM)-dependent methyltransferases.
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PMID:DNA modification by methyltransferases. 777 46

Changes in the pattern of DNA methylation have been a consistent finding in cancer cells. The mostly descriptive nature of these studies and the fact that both hypo- and hypermethylation have been observed at various loci have made it difficult to assess whether these changes are causally involved in the transformation process or whether they reflect the altered physiology of rapidly dividing cancer cells. It is clear, however, that DNA methylation plays an important role in the generation of mutations in human tumors. The high incidence of C-to-T transitions found in the p53 tumor-suppressor gene is attributed to the spontaneous deamination of 5-methylcytosine residues. The multiple observations linking DNA methylation to cancer can be resolved in a model proposing that the high rate of mutation at CpG dinucleotides is due in part to methyltransferase-facilitated deamination. Support for a role of DNA methyltransferase as a mutator enzyme is provided by work with a prokaryotic DNA methyltransferase under S-adenosyl-methionine methyl-donor limiting conditions. Methyl-donor limiting conditions might arise in early stages of tumor development, leading to high rates of methyltransferase-mediated CpG mutagenesis, as seen in human tumors. Such a mechanism is consistent with the frequently reported methionine auxotrophy of cancer cells and with the tumorigenic effects of methyl-deficient diets. Methyl deficiency in tumor cells is also consistent with the commonly observed global hypomethylation of tumor cell DNA, despite normal or even high levels of DNA methyltransferase expression.
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PMID:DNA methylation and cancer. 784 43

We have developed a simple new method that can identify the base methylated by a sequence-specific DNA methyltransferase and have used it to identify the cytosine that is methylated by DsaV methyltransferase (M. DsaV) within its recognition sequence 5'-CCNGG. The method utilizes the fact that exonuclease III of E. coli does not degrade DNA ends with 3' overhangs and cannot hydrolyze a phosphorothioate linkage. DNA duplexes containing phosphorothioate linkages at specific positions were methylated with M. DsaV in the presence of [methyl-3H] S-adenosylmethionine and were subjected to exonuclease III digestion. The pattern of [methyl-3H] dCMP release from the duplexes was consistent with the methylation of the internal cytosine in CCNGG, but not of the outer cytosine. To establish the accuracy of this method, we confirmed the known specificity of EcoRII methyltransferase by the method. We also confirmed the specificity of M. DsaV using an established biochemical method that involves the use of a type IIS restriction enzyme. Methylation of CCWGG (W = A or T) sequences at the internal cytosines is native to E. coli and is not restricted by the modified cytosine restriction (Mcr) systems. Surprisingly, the gene for M. DsaV was significantly restricted by the McrBC system. We interpret this to mean that M. DsaV may occasionally methylate at sequences other than CCNGG or may occasionally methylate the outer cytosine in its recognition sequence.
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PMID:Determination of methylation specificity of DsaV methyltransferase by a simple biochemical method. 787 May 87

The Thermus aquaticus DNA methyltransferase M.Taq I (EC 2.1.1.72) methylates N6 of adenine in the specific double-helical DNA sequence TCGA by transfer of --CH3 from the cofactor S-adenosyl-L-methionine. The x-ray crystal structure at 2.4-A resolution of this enzyme in complex with S-adenosylmethionine shows alpha/beta folding of the polypeptide into two domains of about equal size. They are arranged in the form of a C with a wide cleft suitable to accommodate the DNA substrate. The N-terminal domain is dominated by a nine-stranded beta-sheet; it contains the two conserved segments typical for N-methyltransferases which form a pocket for cofactor binding. The C-terminal domain is formed by four small beta-sheets and alpha-helices. The three-dimensional folding of M.Taq I is similar to that of the cytosine-specific Hha I methyltransferase, where the large beta-sheet in the N-terminal domain contains all conserved segments and the enzymatically functional parts, and the smaller C-terminal domain is less structured.
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PMID:Three-dimensional structure of the adenine-specific DNA methyltransferase M.Taq I in complex with the cofactor S-adenosylmethionine. 797 91


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