Chemical mapping of cytosines enzymatically flipped out of the DNA helix

被引:10
作者
Daujotyte, Dalia [1 ]
Liutkeviciute, Zita [1 ]
Tamulaitis, Gintautas [1 ]
Klimasauskas, Saulius [1 ]
机构
[1] Inst Biotechnol, LT-02241 Vilnius, Lithuania
关键词
D O I
10.1093/nar/gkn200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Haloacetaldehydes can be employed for probing unpaired DNA structures involving cytosine and adenine residues. Using an enzyme that was structurally proven to flip its target cytosine out of the DNA helix, the HhaI DNA methyltransferase (M. HhaI), we demonstrate the suitability of the chloroacetaldehyde modification for mapping extrahelical (flipped-out) cytosine bases in protein-DNA complexes. The generality of this method was verified with two other DNA cytosine-5 methyltransferases, M. AluI and M. SssI, as well as with two restriction endonucleases, R. Ecl18kI and R. PspGI, which represent a novel class of base-flipping enzymes. Our results thus offer a simple and convenient laboratory tool for detection and mapping of flipped-out cytosines in protein-DNA complexes.
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页数:7
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