Control and Dynamical Systems, California Institute of Technology, Pasadena, CA 91125, USA.
Proc Natl Acad Sci U S A. 2011 Oct 4;108(40):E784-93. doi: 10.1073/pnas.1100060108. Epub 2011 Sep 15.
The realization of artificial biochemical reaction networks with unique functionality is one of the main challenges for the development of synthetic biology. Due to the reduced number of components, biochemical circuits constructed in vitro promise to be more amenable to systematic design and quantitative assessment than circuits embedded within living organisms. To make good on that promise, effective methods for composing subsystems into larger systems are needed. Here we used an artificial biochemical oscillator based on in vitro transcription and RNA degradation reactions to drive a variety of "load" processes such as the operation of a DNA-based nanomechanical device ("DNA tweezers") or the production of a functional RNA molecule (an aptamer for malachite green). We implemented several mechanisms for coupling the load processes to the oscillator circuit and compared them based on how much the load affected the frequency and amplitude of the core oscillator, and how much of the load was effectively driven. Based on heuristic insights and computational modeling, an "insulator circuit" was developed, which strongly reduced the detrimental influence of the load on the oscillator circuit. Understanding how to design effective insulation between biochemical subsystems will be critical for the synthesis of larger and more complex systems.
用独特功能的人工生化反应网络的实现是合成生物学发展的主要挑战之一。由于组件数量减少，体外构建的生化电路有望比嵌入生物体内的电路更易于系统设计和定量评估。为了实现这一承诺，需要有效的方法将子系统组合成更大的系统。在这里，我们使用基于体外转录和 RNA 降解反应的人工生化振荡器来驱动各种“负载”过程，例如基于 DNA 的纳米机械装置（“DNA 镊子”）的操作或功能性 RNA 分子（孔雀石绿的适体）的产生。我们实现了将负载过程与振荡器电路耦合的几种机制，并根据负载对核心振荡器的频率和幅度的影响程度以及有效驱动的负载量对它们进行了比较。基于启发式见解和计算建模，开发了一种“绝缘子电路”，它大大降低了负载对振荡器电路的不利影响。了解如何在生化子系统之间设计有效的隔离将是合成更大、更复杂系统的关键。
