Peptide nucleic acid (PNA) probes have been synthesized and targeted to quadruplex DNA. UV-vis and CD spectroscopy reveal that the quadruplex structure of the thrombin binding aptamer (TBA) is disrupted at 37 degrees C by a short PNA probe. The corresponding DNA probe fails to bind to the stable secondary structure at this temperature. Thermal denaturation experiments indicate surprisingly high thermal and thermodynamic stabilities for the PNA-TBA hybrid. Our results point to the nonbonded nucleobase overhangs on the DNA as being responsible for this stability. This "overhang effect" is found for two different PNA-DNA sequences and a variety of different overhang lengths and sequences. The stabilization offered by the overhangs assists the PNA in overcoming the stable secondary structure of the DNA target, an effect which may be significant in the targeting of biological nucleic acids, which will always be much longer than the PNA probe. The ability of PNA to invade a structured DNA target expands its potential utility as an antigene agent or hybridization probe.
DNA guanine (G) quadruplexes are stabilized by an interesting variation of the hydrogen-bonding schemes encountered in nucleic acid duplexes and triplexes. In an attempt to use this mode of molecular recognition, we target a dimeric G-quadruplex formed by the Oxytricha nova telomeric sequence d(G(4)T(4)G(4)) with a peptide nucleic acid (PNA) probe having a homologous rather than complementary sequence. UV-vis and CD spectroscopy reveal that a stable hybrid possessing G-quartets is formed between the PNA and DNA. The four-stranded character of the hybrid and the relative orientation of the strands is determined by fluorescence resonance energy transfer (FRET) experiments. FRET results indicate that (i) the two PNA strands are parallel to each other, (ii) the two DNA strands are parallel to each other, and (iii) the 5'-termini of the DNA strands align with the N-termini of the PNA strands. The resulting PNA(2)-DNA(2) quadruplex shows a preference of Na(+) over Li(+) and displays thermodynamic behavior consistent with alternating PNA and DNA strands in the hybrid. The formation of this novel supramolecular structure demonstrates a new high-affinity DNA recognition mechanism and expands the scope of molecular recognition by PNA.
A series of polyaniline (PANI) oligomers was constructed from monomer units covalently linked to duplex DNA through N-(2-aminoethyl) groups bonded through cytosines. DNA oligomers containing the aniline monomers were treated with horseradish peroxidase (HRP) and H2O2 under conditions known to cause polymerization of aniline. No change in the absorption spectrum of the DNA was observed for samples containing fewer than four contiguous aniline groups. However, for oligomers containing four, five, or six aniline units, treatment with HRP and H2O2 led to the appearance of absorption features characteristic of the conducting "proton doped" emeraldine oxidation state of PANI. Molecular modeling shows that the DNA is distorted in the region of the PANI, but flanking regions of the DNA maintain their B-form structure. These findings provide a method to exploit the self-recognition, self-assembly, and sequence programmability of DNA for the formation of conducting polymers.
A new class of materials was prepared from aniline-containing oligomers that are covalently linked to the nucleobases of duplex DNA. Oligomers composed of repeating aniline (PANI) or 4-aminobiphenyl (PAB) units having the properties of conducting polymers conjoined to the DNA were prepared by the reaction of horseradish peroxidase and H2O2 with DNA having the appropriate monomers aligned within the major groove. These oligomers exhibit the spectral and chemical properties typical of para-linked polyanilines. This method of preparation enables utilization of the unique self-recognizing properties and sequence programmability of DNA to create tailored oligomers. This ability was demonstrated experimentally by preparation of PAB oligomers from alternating benzene and aniline monomers. Conjoined conducting polymers carrying the sequence information of DNA may have applicability as nanowires.
A guanine-rich PNA dodecamer having the sequence H-G4T4G4-Lys-NH2 (G-PNA) hybridizes with a DNA dodecamer of homologous sequence to form a four-stranded quadruplex (Datta, B.; Schmitt, C.; Armitage, B. A. J. Am. Chem. Soc. 2003, 125, 4111-4118). This report describes quadruplex formation by the PNA alone. UV melting curves and fluorescence resonance energy transfer experiments reveal formation of a multistranded structure stabilized by guanine tetrads. The ion dependency of these structures is analogous to that reported for DNA quadruplexes. Electrospray ionization mass spectrometry indicates that both dimeric and tetrameric quadruplexes are formed by G4-PNA, with the dimeric form being preferred. These results have implications for the use of G-rich PNA for homologous hybridization to G-rich targets in chromosomal DNA and suggest additional applications in assembling quadruplex structures within lipid bilayer environments.
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