Context-Sensitive Cleavage of Folded DNAs by Loop-Targeting bPNAs
Context-Sensitive Cleavage of Folded DNAs by Loop-Targeting bPNAs
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DOI：
10.1021/acs.biochem.0c00362
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发表时间：
2020-07-07
期刊：
影响因子：
2.9
通讯作者：
Bong, Dennis
中科院分区：
文献类型：
--
作者：
Liang, Yufeng;Miao, Shiqin;Bong, Dennis
在本文中，我们通过合成bPNA铁和铜配合物，利用氧化主干裂解作为结合的化学读数，证明了上下文依赖的DNA分子识别。显示铁的低聚亚胺bPNAs。EDTA或coppe中心点菲罗啉位点是设计和天然结构dna的高效化学核酸酶，具有丰富的t单链结构域。切割反应性在很大程度上取决于结构背景，如形式（GGGTTA）(n)的DNA底物所显著证明的那样。已知来自人类端粒的重复序列在平行和反平行g -四重体（G4）拓扑结构之间切换，从钾到钠的缓冲液发生变化：值得注意的是，bpna -铜络合物在钠缓冲液中有效地将长重复序列切割成类似于22个核苷酸的部分，而不是钾缓冲液。我们假设反平行拓扑（Na+）比平行拓扑（K+）优先解理，因为TTA环在反平行（Na+）形式下更容易接近bPNA。通过定量聚合酶链反应，可以在PC3细胞的分离DNA和胞内DNA中观察到bPNA核酸酶处理后类似的离子敏感端粒缩短。活细胞治疗伴随着加速的细胞衰老，正如预期的显著端粒缩短。综上所述，bPNA化学核酸酶的环靶向方法补充了先前针对双链和四链DNA的插入策略。结构敏感环靶向可以区分相似的靶序列，从而将bPNA靶向扩展到简单的oligo-T序列之外。此外，bPNA核酸酶具有细胞膜渗透性，因此可用于靶向天然细胞内底物。此外，这些数据表明，bPNA支架可以作为合成特定核酸结构基序的新结合物的平台。
Herein, we demonstrate context-dependent molecular recognition of DNA by synthetic bPNA iron and copper complexes, using oxidative backbone cleavage as a chemical readout for binding. Oligoethylenimine bPNAs displaying iron. EDTA or coppe center dot phenanthroline sites were found to be efficient chemical nucleases for designed and native structured DNAs with T-rich single-stranded domains. Cleavage reactivity depends strongly on structural context, as strikingly demonstrated with DNA substrates of the form (GGGTTA)(n). This repeat sequence from the human telomere is known to switch between parallel and antiparallel G-quadruplex (G4) topologies with a change from potassium to sodium buffer: notably, bPNA-copper complexes efficiently cleave long repeat sequences into similar to 22-nucleotide portions in sodium, but not potassium, buffer. We hypothesize preferential cleavage of the antiparallel topology (Na+) over the parallel topology (K+) due to the greater accessibility of the TTA loop to bPNA in the antiparallel (Na+) form. Similar ion-sensitive telomere shortening upon treatment with bPNA nucleases can be observed in both isolated and intracellular DNA from PC3 cells by quantitative polymerase chain reaction. Live cell treatment was accompanied by accelerated cellular senescence, as expected for significant telomere shortening. Taken together, the loop-targeting approach of bPNA chemical nucleases complements prior intercalation strategies targeting duplex and quadruplex DNA. Structurally sensitive loop targeting enables discrimination between similar target sequences, thus expanding bPNA targeting beyond simple oligo-T sequences. In addition, bPNA nucleases are cell membrane permeable and therefore may be used to target native intracellular substrates. In addition, these data indicate that bPNA scaffolds can be a platform for new synthetic binders to particular nucleic acid structural motifs.