The process of life involves numerous interactions between folded and unfolded large biological molecules. Additionally, the specific folded shape and changes in the shapes of biological molecules can regulate cell cycle events and important cellular functions. Traditionally, all interactions between molecules were thought to fit together like a lock-and-key. However, recent research has shown that nearly 30% of biological molecules are unfolded and interactions between unfolded biological molecules can be "specific" and "transient". Many of these transient interactions involve phase separation (like oil and water) to form sub-cellular structures responsible for clustering the necessary ingredients necessary for cellular functions required for life and replication. This work aims to understand how water concentration and movement at the surface of these biological molecules can alter their shape, structural fluctuations, and phase separation; thus shedding light on important aspects of cellular control and regulation. The project will also provide unique training opportunities for graduate, undergraduate and high school students that include exchange experiences with collaborators and attendance at conferences and workshops. Diverse undergraduate students will be recruited for research experiences that will enable them to integrate their undergraduate course work into a research environment. High school students from underprivileged backgrounds will be recruited into the University of Florida Summer Science Teaching Program.
The expanding discovery of novel RNAs and their unique multifaceted cellular capabilities has resulted in a rich arena of research interfacing RNA biology, chemistry, and biophysics. Intrinsically disordered proteins (IDPs) have also found a growing appreciation for their role in regulating cellular function through the formation of membraneless sub-nuclear organelles. Specifically, this work utilizes site-directed spin-labeling (SDSL) electron paramagnetic resonance approaches to characterize conformational sampling, dynamics and hydration environments of the glycine riboswitch; a large, dynamic RNA that modulates gene expression and select IDP proteins/peptides. The results will reveal how solution environmental conditions modulate conformational flexibility and hydration environment and clarify details of a model of conformational equilibrium that describes interaptamer interactions upon ligand binding that lead to regulatory function (in this case gene expression). This work will also add to the development of spin-labeling magnetic resonance applications in large RNAs; thus laying the foundation for work that can be utilized in other RNA systems. Investigations on the impact of environment on IDPs will provide molecular level details of how environmental parameters (salt, concentration and other molecules) can modulate hydration, structure and phase separation. An ancillary aspect of this project is to continue to optimize Overhauser dynamic nuclear polarization (ODNP) technologies for investigations of macromolecular hydration environment at molecular concentrations found within membraneless organelles such as the nuclear bodies.
