title: Intercalating nucleic acids containing insertions of 1-O-(1-pyrenylmethyl)glycerol: stabilisation of dsDNA and discrimination of DNA over RNA.
abstract: We have studied hybridisation affinities and fluorescence behaviour of intercalator-modified oligonucleotides. The phosphoramidite of (S)-1-O-(4, 4'-dimethoxytriphenylmethyl)-3-O-(1-pyrenylmethyl)glycerol, an intercalating pseudo-nucleotide (IPN), was synthesised and by standard methods inserted into 7mer and 13mer oligodeoxyribonucleotides (ODNs) to generate intercalating nucleic acids (INAs). INAs showed greatly increased affinity for complementary single-stranded DNA (ssDNA), as determined by a thermal stabilisation of the formed DNA/INA duplex of up to 10.9 degrees C per modification when the IPN was added as a dangling end and up to 6.7 degrees C per modification when the IPN was inserted as a bulge. There was a positive stabilisation effect of the formed DNA/INA duplex on introducing a second IPN in the INA strand, when the two IPNs were separated by at least 1 bp. The effect is more pronounced the larger the separation of the two IPNs. Contrary to the enhanced affinity for ssDNA, the IPNs lower the affinity for complementary single-stranded RNA (ssRNA), giving rise to a difference in melting temperature of up to 25.8 degrees C for two IPN insertions in an RNA/INA duplex when compared with the corresponding DNA/INA duplex. In this way INA is able to discriminate ssDNA over ssRNA with identical sequences. Fluorescence measurements show a stronger interaction of the pyrene moiety with DNA than with RNA, indicating intercalation as the stabilising factor in DNA/INA duplexes.
authors.affiliation: Nucleic Acid Center, Department of Chemistry, University of Southern Denmark, DK-5230 Odense M, Denmark.
authors.forename: Ulf B
authors.initials: UB
authors.lastname: Christensen
authors.forename: Erik B
authors.initials: EB
authors.lastname: Pedersen
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000378:metabolism*; D005990:Glycerol / Q000737:chemistry*; D046911:Macromolecular Substances; D008745:Methylation; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000737:chemistry; D009943:Organophosphorus Compounds / Q000737:chemistry; D011721:Pyrenes / Q000737:chemistry*; D012313:RNA / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature
chemical_list: D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D009943:Organophosphorus Compounds; D011721:Pyrenes; C434331:phosphoramidite; D012313:RNA; D004247:DNA; D005990:Glycerol