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

Abstract

Three chimeric dimer synthons (oeg_t^NHT, oeg_up^NHT and oeg_uh^NHT) 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_up^NHT 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_t^NHT < oeg_up^NHT < oeg_uh^NHT. 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.

Key Words:

• 5-Alkynyl-uracils
• PNA-DNA chimeras
• T₂₀-analogues
• Melting properties
• Nuclease stability

Acknowledgments

Financial supports of OTKA (Hungarian Scientific Research Fund, project no.: T 026472) and NKFP MediChem (project no.: 1/047) are gratefully acknowledged. The authors thank Mrs. Emma Belinszki and Mrs. Mária Baraczka for the valuable technical assistance.

Notes

^†Deceased.