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.
