Unambiguous structure characterization of a DNA-RNA triple helix by 15N- and 13C-filtered NOESY spectroscopy.
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.
van Dongen
M J
MJ
NSR Centre for Molecular Structure, Design, and Synthesis, University of Nijmegen, The Netherlands.
Heus
H A
HA
Wymenga
S S
SS
van der Marel
G A
GA
van Boom
J H
JH
Hilbers
C W
CW
D001483:Base Sequence; D002247:Carbon Isotopes; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009587:Nitrogen Isotopes; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics; D009843:Oligoribonucleotides / Q000737:chemistry / Q000235:genetics; D012313:RNA / Q000737:chemistry* / Q000235:genetics
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D009838:Oligodeoxyribonucleotides; D009843:Oligoribonucleotides; D012313:RNA; D004247:DNA