Probing and Manipulating Biomolecules: From Single Molecules to an Ensemble

蛋白质负责执行生命所需的大多数细胞功能，包括细胞内部和细胞之间及其周围环境的催化和分子识别。天然蛋白质也激发了突变体作为新催化剂的工程设计。提出的研究旨在通过结合单分子和集合方法来增强对蛋白质结构-功能关系的理解。为了实现这一目标，我们引入了一种将理论和实验方法相结合的协同策略。我们计划应用：(i)力测量（DFS）来探测生物分子，以改变特定反应的程度甚至命运；（ii）人工酶的合成和天然酶的选择，这些酶的效率可能通过定向进化而提高（由DFS进行探索）；（iii）结合单分子、集合平均和模拟实验对蛋白质动力学进行平行研究，探索它们与细胞膜的相互作用及其功能之间的相互依存关系。该项目的总体目标是在单分子和集合测量之间架起桥梁，进一步了解蛋白质的结构-能量-景观-功能关系，并探索控制细胞内蛋白质扩散动力学的因素，这些因素在细胞信号事件中起着至关重要的作用。我们的努力最终将为活细胞机制的特定方面提供新的见解。

Proteins are responsible for the execution of most cellular functions required for life, including amongothers catalysis and molecular recognition within and between cells and their surroundings. Naturalproteins also inspire the engineering of mutants as new catalysts. The proposed research is aimed atenhancing the understanding of protein structure-function relationships by combining single moleculeand ensemble approaches. To achieve this goal, we introduce a synergetic strategy whichinterdigitates theoretical and experimental approaches. We plan to apply: (i) force measurements(DFS) to probe biological molecules in order to alter the extent and even the fate of specific reactions;(ii) synthesis of artificial enzymes and selection of natural enzymes whose efficiency might beenhanced by directed evolution (to be probed by DFS); (iii) parallel studies on protein dynamicscombining single molecule, ensemble average and simulation experiments to explore theinterdependence between their interactions with the cell membrane and their function. The generallong-range goal of the project is to bridge between single molecule and ensemble measurements, gainfurther understanding of structure-energy landscape-function relationship in proteins, and explore thefactors controlling protein diffusion dynamics in cells, which play crucial roles in cellular signalingevents. Our effort will ultimately contribute new insights into specific aspects of the machinery of aliving cell.