Ultra-stable, high-bandwidth measurement platform for high-precision studies of rapid conformational dynamics in single biomolecules

生物分子，如蛋白质，DNA和RNA，形成复杂的结构，这些结构是它们正确发挥功能所必需的。了解这些结构如何形成（称为“折叠”）对于理解正常功能和功能障碍（可能导致疾病）非常重要。研究折叠最敏感的方法之一是使用激光镊子抓住单个分子，通过将它们拉开来解开它们的结构。该提案将建立一个用于测量单分子折叠的超稳定系统，该系统可以在长时间（数十分钟）内以原子级分辨率观察非常快速（~1 μs）的事件，提供迄今为止最精确的测量这些分子中的实时结构变化。我们将用它来研究基本问题，如折叠如何作为一个物理过程，是什么决定了观察到的特定行为。更具体地说，我们将研究折叠过程中寿命最短的事件，即难以捉摸的“过渡态”，它们在折叠过程中充当瓶颈，因此主导了动力学。通过过渡态的路径包含了折叠机制的所有关键信息，但很难检测到。我们将测量这些“过渡路径”，不仅是为了了解折叠的基本物理原理，也是为了了解折叠是如何出错的：这种错误折叠是许多不治之症的根源，如阿尔茨海默氏症和ALS，但在分子水平上驱动它的机制仍然知之甚少。此外，我们将使用新仪器来研究尚未解决的生物过程，例如病毒如何改变遗传密码的读取方式，这构成了重要的科学之谜，但在医学上也有潜在的应用。像这样的研究，利用单分子探针令人难以置信的分辨率和灵敏度，是折叠研究中最热门的前沿之一。我们现在有一个非凡的机会，可以在一个与生物学、物理学、化学和医学有关的几十年前的问题上取得重大而持久的进展。如果没有这些仪器，我们将无法追求如此令人兴奋的科学。

Biological molecules like proteins, DNA, and RNA form complex structures that are required for them to function correctly. Understanding how these structures form (known as ‘folding’) is important for understanding both proper function and dysfunction (which may lead to disease). One of the most sensitive ways to study folding is to use laser tweezers to grab onto single molecules and unravel their structure by pulling them apart. This proposal will build an ultra-stable system for measuring the folding of single molecules, one that can observe very rapid (~1 μs) events with atomic-scale resolution over long periods of time (tens of minutes), providing the most precise measurements to date of real-time structural changes in these molecules. We will use it to study fundamental questions such as how folding works as a physical process and what determines the specific behaviour observed. More specifically, we will study the very shortest-lived events during folding, the elusive ‘transition states’ that act as bottlenecks in the process and therefore dominate the dynamics. The path taken through the transition states contains all the critical information about the mechanism of folding, but it is very hard to detect. We will measure these ‘transition paths’ not only to learn about the basic physical principles of folding, but also to understand how folding goes wrong: such misfolding is at the root of many incurable diseases, like Alzheimer’s and ALS, but the mechanisms that drive it at the molecular level remain poorly understood. Furthermore, we will use the new instrument to study still-unsolved biological processes like how viruses can alter the way the genetic code is read, which pose important scientific riddles but also have potential applications in medicine. Studies like these, making use of the incredible resolution and sensitivity of single-molecule probes, are one of the hottest frontiers in folding studies. We have right now a remarkable opportunity to make significant and lasting advances in a decades-old problem with relevance to biology, physics, chemistry, and medicine. Without this instrumentation, we will not be able to pursue such exciting science.