Key Laboratory of Green Chemistry and Technology of Ministry of Education, College of Chemistry, Sichuan University, Chengdu, Sichuan 610064, P. R. China.
Department of Chemistry, Centre for Biotechnology, Brock University, St. Catharines, Ontario L2S 3A1, Canada.
J Am Chem Soc. 2021 Sep 1;143(34):13655-13663. doi: 10.1021/jacs.1c05236. Epub 2021 Aug 18.
Chemical modification is a powerful approach to expand the chemical diversity and functionality of natural DNA. However, when chemically modified oligonucleotides are employed in DNA-based reactions or structures, it becomes quite difficult to predict, understand, and control their kinetics and thermodynamics. To address this challenge, we introduce a rationally designed DNA balance capable of measuring critical thermodynamic and kinetic properties of chemically modified DNA in their native environment. Our DNA balance is operated using the principle of toehold-exchange, where a panel of weight probes were designed by tuning the lengths of forward and reverse toeholds. Once placed on the DNA balance, the chemical modification will be interrogated using the weight probes to determine changes in both Gibbs free energy and hybridization rate constant. Using cyclic-azobenzene (cAB)-modified DNA as a model system, we demonstrated that our DNA balance could not only measure stable chemical modifications, but also solve more challenging issues where unstable chemical modifications and transient isomerization reactions were involved. We anticipate that our DNA balance will find wide uses for measuring important thermodynamic and kinetic parameters for DNA carrying various chemical modifications, as well as for probing transient chemical changes in DNA.
化学修饰是一种扩展天然 DNA 化学多样性和功能性的强大方法。然而，当化学修饰的寡核苷酸被用于基于 DNA 的反应或结构中时，预测、理解和控制它们的动力学和热力学变得相当困难。为了解决这个挑战，我们引入了一种经过合理设计的 DNA 平衡，能够在其天然环境中测量化学修饰 DNA 的关键热力学和动力学特性。我们的 DNA 平衡是通过引发链置换的原理来操作的，其中一组重量探针是通过调整正向和反向引发链的长度来设计的。一旦放在 DNA 平衡上，就可以使用重量探针来检测化学修饰，以确定吉布斯自由能和杂交速率常数的变化。我们使用环偶氮苯 (cAB)-修饰的 DNA 作为模型系统，证明我们的 DNA 平衡不仅可以测量稳定的化学修饰，还可以解决更具挑战性的问题，其中涉及不稳定的化学修饰和瞬态异构化反应。我们预计我们的 DNA 平衡将广泛用于测量携带各种化学修饰的 DNA 的重要热力学和动力学参数，以及探测 DNA 中的瞬态化学变化。
