Chemical & Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland 21218, United States.
Biophysics and Biophysical Chemistry, Johns Hopkins University, Baltimore, Maryland 21218, United States.
ACS Synth Biol. 2023 Nov 17;12(11):3424-3432. doi: 10.1021/acssynbio.3c00452. Epub 2023 Oct 16.
The ability to finely tune reaction rates and binding energies between components has made DNA strand displacement circuits promising candidates to replicate the complex regulatory functions of biological reaction networks. However, these circuits often lack crucial properties, such as signal turnover and the ability to transiently respond to successive input signals that require the continuous input of chemical energy. Here, we introduce a method for providing such energy to strand displacement networks in a controlled fashion: an engineered DNA helicase, Rep-X, that transiently dehybridizes specific DNA complexes, enabling the strands in the complex to participate in downstream hybridization or strand displacement reactions. We demonstrate how this process can direct the formation of specific metastable structures by design and that this dehybridization process can be controlled by DNA strand displacement reactions that effectively protect and deprotect a double-stranded complex from unwinding by Rep-X. These findings can guide the design of active DNA strand displacement regulatory networks, in which sustained dynamical behavior is fueled by helicase-regulated unwinding.
精细调节组分之间的反应速率和结合能的能力使 DNA 链置换电路成为复制生物反应网络复杂调控功能的有前途的候选者。然而，这些电路通常缺乏关键特性，例如信号转换和暂时响应需要连续输入化学能量的连续输入信号的能力。在这里，我们介绍了一种以受控方式向链置换网络提供这种能量的方法：一种工程化的 DNA 解旋酶 Rep-X，它瞬时解杂交特定的 DNA 复合物，使复合物中的链能够参与下游杂交或链置换反应。我们展示了如何通过设计来指导特定亚稳态结构的形成，并且该解杂交过程可以通过 DNA 链置换反应来控制，该反应可以有效地保护和解保护双链复合物免受 Rep-X 的解旋。这些发现可以指导活性 DNA 链置换调控网络的设计，其中持续的动力学行为由解旋酶调节的解旋来提供动力。
