Single-Molecule Methods for Interfacial Dynamics

在这个由化学部门化学测量和成像项目资助的研究项目中，科罗拉多大学博尔德分校的Daniel K Schwartz教授和他的团队将开发基于全内反射荧光（TIRF）显微镜的单分子（SM）成像方法，包括动态SM共振能量转移（RET）方法，以探测纳米尺度的分子环境和分子构象。同时观察固/液界面的动态分子行为，包括吸附、解吸、分子间缔合（即聚集）和界面扩散。正在进行的具体实验将(1)将分子构象直接与界面动力学联系起来，(2)明确探测近表面环境中分子间结合（例如聚类，聚集）的动力学。一些新方法涉及移动供体标记探针和受体标记分子基质之间的分子间RET。这些方法提供了关于界面环境如何影响吸附、表面停留时间、界面迁移率和分子识别动力学的直接信息。其他方法使用分子内RET来探测多肽和寡核苷酸的构象变化，并将这些构象变化与动态界面行为联系起来。作为本研究的一部分开发的方法代表了将在研究界广泛应用于研究材料界面的新技术，特别是软材料（如聚合物）的“湿”界面。对这些现象的更好理解将导致生物材料、药物分离和配方、基于分子识别的医学诊断和自组装纳米材料的改进。表面和生物分子之间的相互作用与生物技术和人类健康有着广泛的相关性，影响着医学诊断、治疗性蛋白质稳定性、疫苗效力/安全性和生物材料设计。特别是，这里开发的方法将广泛适用于生物材料的表征，允许对表面化学影响分子构象和横向相互作用的方式的新水平的理解。参与该项目的研究人员将获得在未来几十年预计处于科学技术前沿的多学科领域的专业知识，包括单分子显微镜和自组装分子系统。此外，通过参与各种教育和推广项目，研究人员将与学生、教师和其他社区成员分享这个项目。

In this research project funded by the Chemical Measurements and Imaging program in the Chemistry Division, Prof. Daniel K Schwartz and his group at the University of Colorado at Boulder will develop single-molecule (SM) imaging methods, based on total internal reflection fluorescence (TIRF) microscopy, including dynamic SM resonance energy transfer (RET) methods, to probe the nm-scale molecular environment and molecular conformation, while simultaneously observing dynamic molecular behavior at the solid/liquid interface, including adsorption, desorption, intermolecular association (i.e., aggregation), and interfacial diffusion. Specific experiments are being performed that will (1) link molecular conformation directly to interfacial dynamics, and (2) explicitly probe the dynamics of intermolecular association (e.g. clustering, aggregation) in the near-surface environment. Some of the new methods involve intermolecular RET between mobile donor-labeled probes and an acceptor-labeled molecular matrix. These methods provide direct information about the ways in which the interfacial environment affects adsorption, surface residence time, interfacial mobility, and molecular recognition kinetics. Other methods use intramolecular RET to probe conformational changes of polypeptides and oligonucleotides, and to correlate these conformational changes with dynamic interfacial behavior. The methods developed as part of this research represent enabling new techniques that will be broadly used in the research community to study materials interfaces, especially "wet" interfaces of soft materials such as polymers. An improved understanding of these phenomena will lead to improvements in biomaterials, pharmaceutical separations and formulation, medical diagnostics based on molecular recognition, and self-assembled nanomaterials. Interactions between surfaces and biomolecules have widespread relevance to biotechnology and human health, affecting medical diagnostics, therapeutic protein stability, vaccine efficacy/safety, and biomaterial design. In particular, the methods developed here will be widely-applicable to biomaterials characterization, permitting a new level of understanding of the ways in which surface chemistry influences molecular conformation and lateral interactions. The researchers involved in this project will gain expertise in multidisciplinary areas projected to be at the forefront of science and technology in the coming decades, including single-molecule microscopy and self-assembled molecular systems. Also, through their participation in a variety of educational and outreach programs, the researchers will share this project with students, teachers, and other community members.