Sequence-specific methyltransferase-induced labeling of DNA (SMILing DNA).
نویسندگان
چکیده
A new concept for sequence-specific labeling of DNA by using chemically modified cofactors for DNA methyltransferases is presented. Replacement of the amino acid side chain of the natural cofactor S-adenosyl-L-methionine with an aziridine group leads to a cofactor suitable for DNA methyltransferase-catalyzed sequence-specific coupling with DNA. Sequence-specifically fluorescently labeled plasmid DNA was obtained by using the DNA methyltransferase from Thermus aquaticus (M.TaqI) as catalyst and attaching a fluorophore to the aziridine cofactor. First results suggest that all classes of DNA methyltransferases with different recognition sequences can be used. In addition, this novel method for DNA labeling should be applicable to a wide variety of reporter groups.
منابع مشابه
Sequence-specific labeling of nucleic acids and proteins with methyltransferases and cofactor analogues.
S-Adenosyl-l-methionine (AdoMet or SAM)-dependent methyltransferases (MTase) catalyze the transfer of the activated methyl group from AdoMet to specific positions in DNA, RNA, proteins and small biomolecules. This natural methylation reaction can be expanded to a wide variety of alkylation reactions using synthetic cofactor analogues. Replacement of the reactive sulfonium center of AdoMet with ...
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DNA methyltransferases catalyse the transfer of a methyl group from the ubiquitous cofactor S-adenosyl-L-methionine (AdoMet) onto specific target sites on DNA and play important roles in organisms from bacteria to humans. AdoMet analogs with extended propargylic side chains have been chemically produced for methyltransferase-directed transfer of activated groups (mTAG) onto DNA, although the ef...
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The study of changes in genome-wide levels of DNA methylation has become a key focus for understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA methylation, falling into two categories: gene-specific and genome-wide. Genome-wide methylation analysis is best performed by DNA hydrolysis followed by HPLC; however, it requires access to an HPLC machine, which...
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Bending of DNA induced by M.Msp I, one of the m5C-DNA methyltransferases, has been investigated using circular permutation analysis. The M.Msp I MTase induced sharp bends in DNA containing its recognition sequence 5'-CCGG-3'which was estimated to be 142+/-4 degrees and 132+/-4 degrees for circularly permuted DNA fragments of 127 and 1459 bp respectively. The bend centre was found to be asymmetr...
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عنوان ژورنال:
- Chembiochem : a European journal of chemical biology
دوره 5 3 شماره
صفحات -
تاریخ انتشار 2004