Department of Physics, Center for Nanoscience, Ludwig-Maximilians-Universität München, Amalienstraße 54, 80799 Munich, Germany.
Department of Physics and Astronomy, McMaster University, 1280 Main Street West, Hamilton, Ontario L8S4M1, Canada.
J Am Chem Soc. 2024 Apr 3;146(13):8887-8894. doi: 10.1021/jacs.3c10813. Epub 2024 Mar 19.
Templated ligation offers an efficient approach to replicate long strands in an RNA world. The 2',3'-cyclic phosphate (>P) is a prebiotically available activation that also forms during RNA hydrolysis. Using gel electrophoresis and high-performance liquid chromatography, we found that the templated ligation of RNA with >P proceeds in simple low-salt aqueous solutions with 1 mM MgCl under alkaline pH ranging from 9 to 11 and temperatures from -20 to 25 °C. No additional catalysts were required. In contrast to previous reports, we found an increase in the number of canonical linkages to 50%. The reaction proceeds in a sequence-specific manner, with an experimentally determined ligation fidelity of 82% at the 3' end and 91% at the 5' end of the ligation site. With splinted oligomers, five ligations created a 96-mer strand, demonstrating a pathway for the ribozyme assembly. Due to the low salt requirements, the ligation conditions will be compatible with strand separation. Templated ligation mediated by 2',3'-cyclic phosphate in alkaline conditions therefore offers a performant replication and elongation reaction for RNA on early Earth.
模板连接提供了一种有效的方法来复制 RNA 世界中的长链。2',3'-环磷酸酯(>P)是一种可预先获得的激活物,也会在 RNA 水解过程中形成。通过凝胶电泳和高效液相色谱分析,我们发现> P 模板连接 RNA 可在简单的低盐水性溶液中,在碱性 pH 值 9 到 11 范围内和-20 到 25°C 的温度下进行,无需额外的催化剂。与之前的报告不同,我们发现增加了 50%的典型连接数。该反应具有序列特异性,在 3'末端的连接保真度为 82%,在连接位点的 5'末端为 91%。对于带有间隔寡核苷酸的反应,五次连接可产生 96 -mer 链,证明了核酶组装的途径。由于盐浓度要求低,连接条件将与链分离兼容。因此,在碱性条件下由 2',3'-环磷酸介导的模板连接为早期地球上的 RNA 提供了一种高效的复制和延伸反应。