PNA-DNA Chimeras Containing 5-Alkynyl-pyrimidine PNA Units. Synthesis, Binding Properties, and Enzymatic Stability

Abstract

Three chimeric dimer synthons (oeg_tNHT, oeg_upNHT and oeg_uhNHT) containing thymine (t), 5-(l-propynyl)-uracil (up) and 5-(1-hexyn-1-yl)-uracil (uh) PNA units with N-(2-hydroxyethyl)glycine (oeg) backbone were synthesized in solution and incorporated into T20 oligonucleotide analogues, using standard P-amidite chemistry. Insertion of dimer blocks led to destabilization of duplexes with dA20 target. The smallest T m drops were found for chimeras containing oeg_upNHT dimers. Incorporation of the chimeric synthons into the 3′-end of T20 brought about growing resistance to 3′-exonucleolytic (SV PDE) cleavage in the order of oeg_tNHT < oeg_upNHT < oeg_uhNHT. Due to different endonuclease activities of 3′- and 5′-exonucleases applied, placing of five consecutive dimers at the 5′-terminus resulted in a relatively smaller, but also side-chain dependent, stabilization towards the hydrolysis by 5′-exonuclease (BS PDE). Neither exonucleases (SV and BS PDE) nor an endonuclease (Nuclease P1) could hydrolyse the unnatural phosphodiester bond linking the 3′-OH of thymidine to the terminal OH of N-(2-hydroxyethyl)glycine PNA backbone.