科研之友
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摘要
The guanine quadruplex (G-quadruplex) is a highly stable secondary structure that forms in G-rich repeats of DNA, which can interfere with DNA processes, including DNA replication and transcription. We showed previously that short guanine-rich peptide nucleic acids (PNAs) can form highly stable hybrid quadruplexes with DNA. We hypothesized that such structures would provide a stronger block to polymerase extension on G-rich templates than a native DNA homoquadruplex because of the greater thermodynamic stability of the PNA-DNA hybrid structures. To test this, we analyzed the DNA primer extension activity of polymerase eta, a translesion polymerase implicated in synthesis past G-quadruplex blocks, on DNA templates containing guanine repeats. We observed a PNA concentration-dependent decrease in the level of polymerase eta extension to the end of the template and an increase in the level of polymerase eta inhibition at the sequence prior to the G-rich repeats. In contrast, the addition of a complementary C-rich PNA that hybridizes to the G-rich repeats by Watson-Crick base pairing led to a decrease in the level of polymerase inhibition and an increase in the level of full-length extension products. The G-quadruplex-forming PNA exhibited inhibition (IC50 = 16.2 +/- 3.3 nM) of polymerase eta DNA synthesis on the G-rich templates stronger than that of the established G-quadruplex-stabilizing ligand BRACO-19 (IC50 = 42.5 +/- 4.8 nM). Our results indicate that homologous PNA targeting of G-rich sequences creates stable PNA-DNA heteroquadruplexes that inhibit polymerase eta extension more effectively than a DNA homoquadruplex. The implications of these results for the potential development of homologous PNAs as therapeutics for halting proliferating cancer cells are discussed.
- 出版日期2014-8-19
