[JACS] DNA in a Nano-Sized Frame Successfully Observed to Respond to Enzymatic Modification

January 15, 2010

Kyoto University's Institute for Integrated Cell-Material Sciences (iCeMS) Assoc. Prof. Masayuki Endo and Prof. Hiroshi Sugiyama have successfully observed the DNA methylation enzyme EcoRI methyltransferase (M.EcoRI) selectively binding to a particular DNA strand in a specially fabricated DNA origami frame.

Their findings have been published in the online edition of the Journal of the American Chemical Society.


Assoc. Prof. Masayuki Endo

Prof. Hiroshi Sugiyama

Extenal Link Regulation of DNA Methylation Using Different Tensions of Double Strands Constructed in a Defined DNA Nanostructure
Masayuki Endo,1,2 Yousuke Katsuda,2 Kumi Hidaka,2 and Hiroshi Sugiyama1,2,3
  1. Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University
  2. Department of Chemistry, Graduate School of Science, Kyoto University
  3. Japan Science and Technology Corporation (JST), CREST


Journal of the American Chemical Society*, published online Jan 15, 2010.
doi:10.1016/ja907649w

[Abstract]
A novel strategy for regulation of an enzymatic DNA modification reaction has been developed by employing a designed nanoscale DNA scaffold. DNA modification using enzymes often requires bending of specific DNA strands to facilitate the reaction. The DNA methylation enzyme EcoRI methyltransferase (M.EcoRI) bends double helix DNA by 55°−59° during the reaction with flipping out of the second adenine in the GAATTC sequence as the methyl transfer reaction proceeds. In this study, two different double helical tensions, tense and relaxed states of double helices, were created to control the methyl transfer reaction of M.EcoRI and examine the structural effect on the methylation. We designed and prepared a two-dimensional (2D) DNA scaffold named the "DNA frame" using the DNA origami method that accommodates two different lengths of the double-strand DNA fragments, a tense 64mer double strand and a relaxed 74mer double strand. Fast-scanning atomic force microscope (AFM) imaging revealed the different dynamic movement of the double-strand DNAs and complexes of M.EcoRI with 64mer and 74mer double-strand DNAs. After treatment of the double strands in the DNA frame with M.EcoRI and the subsequent digestion with restriction enzyme EcoRI (R.EcoRI), AFM analysis revealed that the 74mer double-strand DNA was not effectively cleaved compared with the 64mer double-strand DNA, indicating that the methylation preferentially occurred in the relaxed 74mer double-strand DNA compared with that in the tense 64mer double strand. Biochemical analysis of the methylation and specific digestion using a real-time PCR also supported the above results. These results indicate the importance of the structural flexibility for bending of the duplex DNA during the methyl transfer reaction with M.EcoRI. Therefore, the DNA methylation can be regulated using the structurally controlled double-strand DNAs constructed in the DNA frame nanostructure.

*Impact factor: 8.091
  (7th/127 in Multidisciplinary Chemistry, Thomson Reuters 2008 Journal Citation Reports)


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