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Presented By: Department of Chemistry

Kevin Welsher

Kevin Welsher (Duke University)

The biochemical environment of the cellular interior is extremely complex and plays a large role in the function of all biomolecules necessary to life. In this talk, I will describe two ways our group is trying to understand how the cellular environment affects biomolecule function using single molecule and biophysical tools. In the first part, I will discuss how single molecule studies have been impeded by the rapid 3D diffusion of single molecules in live cells, precluding long-duration and high-temporal resolution measurement. To overcome this hurdle, we have developed 3D single-molecule active real-time tracking (3D-SMART) which enables active feedback tracking of rapidly diffusing and lowly emitting fluorescent particles, from single virus-like particles and quantum dots in water, all the way down to single proteins and nucleic acids in viscous solution. 3D-SMART represents a new path toward high resolution single molecule spectroscopy on untethered molecules. In the second half of the talk, I will discuss our efforts to study the structure of the cytoplasm itself, starting with intracellular water. Despite its fundamental nature, direct visualization of subcellular solvation heterogeneity has remained elusive. To explore this question, we have demonstrated a vibrational-shift imaging approach to probe solvation at the microscopic level by combining spectral-focusing hyperspectral stimulated Raman scattering (hsSRS) with an environmentally-sensitive nitrile probe. When applied to quantitatively measure the spatial variation of solvation in live cells, this new method reveals significantly reduced solvation in the cytoplasm compared to the nuclear compartment and bulk water! This work sheds light on heterogenous solvation at the subcellular level and opens up new avenues to explore solvation variance in complex systems.
Kevin Welsher (Duke University)

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