Single Molecule Studies of Biomolecules

单分子荧光检测方法可以提供对蛋白质的随机动力学和结构分布的洞察。在这里，我们建议开发生化和光谱工具，以促进单分子方法在蛋白质结构和功能研究中的应用。这些方法包括：(1)用于荧光研究和表面固定的带有适当连接物和染料的蛋白质选择性衍生化方法；(2)用于单分子荧光检测的仪器和数据分析工具/方法。这些方法将被应用于三个明确的生化系统：(I)胰蛋白酶对多肽和蛋白质的酶解；(Ii)去肌红蛋白的折叠；以及(Iii)膜蛋白与C5a受体的结合和扩散的研究。

Single molecule fluorescence detection methods can provide insights into the stochastic dynamics and structural distribution of proteins. Here we propose to develop both biochemical and spectroscopic tools to facilitate the application of single molecule methods to the study of protein structure and function. These include: (i) methods for selective derivatization of proteins with appropriate linkers and dyes for fluorescence studies and surface immobilization; and (ii) instrumentation and data analysis tools/methods for single molecule fluorescence detection. These methods will be applied to three well-defined biochemical systems: (i) the enzymatic hydrolysis of peptides and proteins by trysin; (ii) the folding of apomyoglobin; and (iii) studies of membrane protein association and diffusion with C5a receptor.