Wen Junlin, Zhang Chunlei, Chen Xue, Dai Ziwen, Li Mengting, Ma Wenjian, Yam ChiYung, Huang Xiaoluo, Xiong Chenghe, Mei Hui
Shenzhen Key Laboratory of Synthetic Genomics, Guangdong Provincial Key Laboratory of Synthetic Genomics, State Key Laboratory of Quantitative Synthetic Biology, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China.
Gingko Biotech Limited, Hong Kong SAR, 000000, China.
Commun Chem. 2025 May 13;8(1):144. doi: 10.1038/s42004-025-01545-8.
End capping of oligonucleotides by modified nucleotides is essential for boosting resistance to 3' exonuclease degradation, thereby enhancing their stability and therapeutic efficacy in vivo. However, the rationale behind these modifications remains unclear. In this study, we designed a novel nucleic acid analog, eTNA, by replacing deoxyribose with the α-D-erythrofuranosyl moiety. As an epimer of TNA (threose nucleic acid), it combines structural features from inverted-dT and TNA, both known for enhancing resistance against 3'-exonucleases. On top of this, we systematically investigated the stability of a series of oligonucleotides capped with inverted-dT, TNA and eTNA at the 5'-, 3'-, or both ends. The structural differences between eTNA and natural dT help to understand how the sugar ring's conformation and rigidity affect duplex stability and exonuclease resistance. Our experimental and theoretical results show that the modified furanose affects the binding positions of terminal nucleotides in the phosphodiesterase active site, preventing phosphodiester hydrolysis. Our mechanistic study should benefit future therapeutic oligonucleotide design with end capping.
通过修饰核苷酸对寡核苷酸进行封端对于提高其对 3' 核酸外切酶降解的抗性至关重要，从而增强其在体内的稳定性和治疗效果。然而，这些修饰背后的原理仍不清楚。在本研究中，我们通过用α-D-赤藓糖呋喃糖部分取代脱氧核糖设计了一种新型核酸类似物 eTNA。作为 TNA（苏糖核酸）的差向异构体，它结合了来自反向 dT 和 TNA 的结构特征，这两者都以增强对 3'-核酸外切酶的抗性而闻名。除此之外，我们系统地研究了一系列在 5'-、3'- 或两端用反向 dT、TNA 和 eTNA 封端的寡核苷酸的稳定性。eTNA 与天然 dT 之间的结构差异有助于理解糖环的构象和刚性如何影响双链稳定性和核酸外切酶抗性。我们的实验和理论结果表明，修饰的呋喃糖会影响磷酸二酯酶活性位点中末端核苷酸的结合位置，从而防止磷酸二酯水解。我们的机理研究应该会有益于未来带有封端的治疗性寡核苷酸设计。
