Department of Food Processing Technology and Management

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    A NOVEL SECONDARY DNA BINDING SITE IN HUMAN TOPOISOMERASE I UNRAVELLED BY USING A 2D DNA ORIGAMI PLATFORM
    (ACS Publications, 2010-09-09) Ramesh, Subramani; Sissel, Juul; Alexandru, Rotaru; Felicie F, Andersen; Kurt V., Gothelf; Wael, Mamdouh; Flemming, Besenbacher; Mingdong, Dong; Birgitta R, Knudsen
    The biologically and clinically important nuclear enzyme human topoisomerase I relaxes both positively and negatively supercoiled DNA and binds consequently DNA with supercoils of positive or negative sign with a strong preference over relaxed DNA. One scheme to explain this preference relies on the existence of a secondary DNA binding site in the enzyme facilitating binding to DNA nodes characteristic for plectonemic DNA. Here we demonstrate the ability of human topoisomerase I to induce formation of DNA synapses at protein containing nodes or filaments using atomic force microscopy imaging. By means of a two-dimensional (2D) DNA origami platform, we monitor the interactions between a single human topoisomerase I covalently bound to one DNA fragment and a second DNA fragment protruding from the DNA origami. This novel single molecule origami-based detection scheme provides direct evidence for the existence of a secondary DNA interaction site in human topoisomerase I and lends further credence to the theory of two distinct DNA interaction sites in human topoisomerase I, possibly facilitating binding to DNA nodes characteristic for plectonemic supercoils.
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    SINGLE-MOLECULE CHEMICAL REACTIONS ON DNA ORIGAMI
    (Nature Nanotechnology, 2010-02-28) Niels V, Voigt; Thomas, Tørring; Alexandru, Rotaru; Mikkel F, Jacobsen; Jens B, Ravnsbæk; Ramesh, Subramani; Wael, Mamdouh; Jørgen, Kjems; Andriy, Mokhir; Flemming, Besenbacher; Kurt, Vesterager Gothelf
    DNA nanotechnology1,2 and particularly DNA origami3, in which long, single-stranded DNA molecules are folded into predetermined shapes, can be used to form complex self-assembled nanostructures4,5,6,7,8,9,10. Although DNA itself has limited chemical, optical or electronic functionality, DNA nanostructures can serve as templates for building materials with new functional properties. Relatively large nanocomponents such as nanoparticles and biomolecules can also be integrated into DNA nanostructures and imaged11,12,13. Here, we show that chemical reactions with single molecules can be performed and imaged at a local position on a DNA origami scaffold by atomic force microscopy. The high yields and chemoselectivities of successive cleavage and bond-forming reactions observed in these experiments demonstrate the feasibility of post-assembly chemical modification of DNA nanostructures and their potential use as locally addressable solid supports.