首页> 外文期刊>Biochemistry >Hybridization of G?Quadruplex-Forming Peptide Nucleic Acids to Guanine-Rich DNA Templates Inhibits DNA Polymerase η Extension
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Hybridization of G?Quadruplex-Forming Peptide Nucleic Acids to Guanine-Rich DNA Templates Inhibits DNA Polymerase η Extension

机译:G?四链体形成肽核酸与鸟嘌呤丰富的DNA模板的杂交抑制DNA聚合酶η延伸。

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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 η, a translesion polymerase implicated in synthesis past G-quadruplex blocks, on DNA templates containing guanine repeats. We observed a PNA concentrationdependent decrease in the level of polymerase η extension to the end of the template and an increase in the level of polymerase η 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 (IC_(50) = 16.2 ± 3.3 nM) of polymerase η DNA synthesis on the G-rich templates stronger than that of the established G-quadruplex-stabilizing ligand BRACO-19 (IC_(50) = 42.5 ± 4.8 nM). Our results indicate that homologous PNA targeting of G-rich sequences creates stable PNA?DNA heteroquadruplexes that inhibit polymerase η 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.
机译:鸟嘌呤四联体(G-quadruplex)是高度稳定的二级结构,可在富含G的DNA重复序列中形成,可干扰DNA的过程,包括DNA复制和转录。我们以前已经证明,富含鸟嘌呤的短肽核酸(PNA)可以与DNA形成高度稳定的杂交四链体。我们推测,由于PNA?DNA杂合结构具有更高的热力学稳定性,与天然DNA同四聚体相比,此类结构在富G模板上可提供更强的聚合酶延伸阻断作用。为了测试这一点,我们在包含鸟嘌呤重复序列的DNA模板上分析了聚合酶η的DNA引物延伸活性,聚合酶η通过G-四链体嵌段参与合成。我们观察到在富G重复之前的序列中,聚合酶η延伸至模板末端的水平随PNA浓度的降低而降低,而聚合酶η抑制水平的升高。相反,通过Watson?Crick碱基配对添加与C富集重复序列杂交的C富集互补PNA导致聚合酶抑制水平的降低和全长延伸产物水平的提高。形成G-四链体的PNA对富G模板的聚合酶ηDNA合成的抑制作用(IC_(50)= 16.2±3.3 nM)比已建立的G-四链体稳定配体BRACO-19(IC_(50 )= 42.5±4.8 nM)。我们的结果表明,同源的PNA靶向富G序列可产生稳定的PNAαDNA异四聚体,该复合物比DNA同四聚体更有效地抑制聚合酶η延伸。讨论了这些结果对于同源PNAs潜在发展作为停止增殖癌细胞的治疗方法的意义。

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