Polyanionic Carboxyethyl Peptide Nucleic Acids (ce-PNAs): Synthesis and DNA Binding.

New polyanionic modifications of polyamide nucleic acid mimics were obtained. Thymine decamers were synthesized from respective chiral α- and γ-monomers, and their enantiomeric purity was assessed. Here, we present the decamer synthesis, purification and characterization by MALDI-TOF mass spectrometry and an investigation of the hybridization properties of the decamers. We show that the modified γ-S-carboxyethyl-T10 PNA forms a stable triplex with polyadenine DNA

Schematic representation of PNA and DNA fragments.

<p>(A) DNA fragment; (B) <i>aeg</i>-, γ- and α-<i>ce</i>-PNAs; (C) Starting thymine monomers <b>4</b> and <b>5</b> and their conformers, which coexist in solution due to the reduced rotation along the N^<b>tert</b>-C(O)-bond.</p

Chromatographic profiles of the PNA monomer derivatives (diastereomers).

<p>(A) Mixture of diastereomers <b>7a</b> and <b>7b</b>; (B) Diastereomer <b>7a</b> (Conditions: Luna column, 5 μm, CN 100A, elution system: heptane/isopropanol (88/12), flow rate: 1 mL/min); (C) Mixture of diastereomers <b>8a</b> and <b>8b</b>; (D) Diastereomer <b>8a</b> (Conditions: Separon SGX RP-S C18 column, 5 μm, 4x150 mm, elution system: acetonitrile/water (1/1), flow rate: 1 mL/min); R = -CH₂CH₂COOBn.</p

Chromatographic profiles of the PNA monomers (enantiomers).

<p>(A) Mixture of <b>4a</b> and <b>4b</b> (<i>S</i>- and <i>R</i>- enantiomers of the γ-monomer); (B) <i>S</i>-entantiomer of the γ-monomer, <b>4a</b> (Conditions: Diasphere column 110-Chirasel-E-PA, 7 μm; elution system: MeOH/CH₃COOH = 96/4 (v/v); flow rate: 1 mL/min; UV-detection at 254 nm; temperature: 20°C); (C) Mixture of <b>5a</b> and <b>5b</b> (<i>R</i>- and <i>S</i>-enantiomers of the α-monomer); (D) <i>S</i>-entantiomer of the α-monomer, <b>5a</b> (Conditions: Diasphere column 110-Chirasel-E-PA, 7 μm; elution system: MeOH/CH₃COOH/TEA = 100/0.1/0.1 (v/v/v); flow rate: 0.5 mL/min; UV-detection at 254 nm; temperature: 20°C); R = CH₂CH₂COOBn.</p

J-Plots and CD spectra of γ-<i>ce</i>-H-(T₁₀)-Gly-NH₂ (2)/dA₁₀.

<p>(A) Absorbance of the γ-<i>ce</i>-H-(T₁₀)-Gly-OH (<b>2</b>)/dA₁₀ mixtures with various PNA/DNA ratios at 260 nm; (B) Absorbance of the mixtures at 275 nm; (C) Molar ellipticity at 263 nm. Conditions: 10 mM Na₂HPO₄ (pH 7.4), 140 mM KCl, 5 mM MgCl₂. The summed PNA and DNA concentration was 7 μM at each ratio; (D) CD spectra of the triplex. The molar ellipticity is given per 1 nucleotide in a strand. Conditions: 10 mM Na₂HPO₄ (pH 5 or 7.4), 140 mM KCl, 5 mM MgCl₂. The triplex concentration was 1.8 μM.</p

UV-melting and CD spectra of PNA/DNA and DNA/DNA complexes.

<p>(A) Thermal difference spectra of the complexes; (B) Thermal denaturation profiles of the complexes; (C) CD spectra of the single-stranded oligonucleotides and PNAs. The molar ellipticity was calculated per 1 nucleotide. (D) CD spectra of the complexes. Conditions: 10 mM Na₂HPO₄ (pH 7.4), 140 mM KCl, 5 mM MgCl₂. The concentration of each PNA or oligonucleotide was 2.5 μM. The CD spectra were measured at 15°C.</p