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  • Title: Unambiguous structure characterization of a DNA-RNA triple helix by 15N- and 13C-filtered NOESY spectroscopy.
    Author: van Dongen MJ, Heus HA, Wymenga SS, van der Marel GA, van Boom JH, Hilbers CW.
    Journal: Biochemistry; 1996 Feb 13; 35(6):1733-9. PubMed ID: 8639652.
    Abstract:
    DNA.DNA*RNA triple helices of the pyrimidine.purine*pyrimidine motif (where . indicates Watson-Crick pairing and * indicates Hoogsteen pairing) appear to be very stable, which has important implications for the development of novel antisense strategies. Here we present the first structural NMR studies on such a system, composed of a DNA hairpin with a homopurine-homopyrimidine stem sequence and a single-stranded RNA oligonucleotide containing exclusively pyrimidine residues. In these investigations an unlabeled DNA hairpin and a uniformly 13C/15N-enriched RNA oligonucleotide were utilized in combination with X-edited 1H NMR spectroscopy. Improved 15N (omega 2) filtered NOESY and 13C (omega 1) filtered NOESY are presented by which we were able to differentiate between intrastrand, i.e., DNA-DNA and RNA-RNA, and interstrand, i.e., DNA-RNA, NOE contacts. It is unambiguously established that the complex forms a right-handed triple helix, with the RNA strand situated in the major groove of the Watson-Crick stem of the hairpin. The interaction is stabilized by the formation of Hoogsteen-type base pairs between the RNA strand and the purine strand of the DNA. These strands run parallel to each other. The characterization of the DNA-RNA triple helix structure described here shows that this type of experiment forms a valuable instrument in the structure determination of bimolecular systems of nucleic acids.
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