ZOOM LINK:   https://gc-cuny.zoom.us/j/4954048198?pwd=eVlkMFdHcjV6d3pkYzB4V2VtbHJGdz…
Meeting ID: 495 404 8198

Raghavendar Sanganna Gari, Postdoctoral Associate, Anesthesiology, Weill Cornell Medicine, will give a talk on "Correlation of membrane protein dynamics with function."

ABSTRACT

Membrane proteins (MPs) embedded in a lipid bilayer regulate the flow of ions, nucleotides, small molecules, lipids, and proteins across or into the membranes in all cells. Despite their crucial role in health and disease, the molecular mechanisms of many MPs remains elusive. This is due to the challenges in obtaining structural dynamics in a lipid bilayer environment, and correlating dynamics with structure, and function. In this work, we attempted to bridge the gap between structure, function, and dynamics of MPs using high-speed atomic force microscopy (HS-AFM) technique and the E. coli β-barrel outer membrane protein OmpG as a model system. HS-AFM provides unprecedented real-space and real-time visualization of biological molecules in aqueous environment and at ambient temperature. OmpG is imbued with both structural and functional simplicity, where the motions of a single loop (loop-6) are supposed to directly open and close the ion conducting pore, transitions which can be characterized with single-channel recordings. X-ray structures of OmpG revealed that at neutral pH, the channel (barrel) is open, while at pH 5.6 access to the barrel is blocked by loop-6, which folds into the barrel lumen in a lid-like manner. However, a direct correlation between structural conformations and channel functional states was not possible because of the lack of a time-resolved single-molecule structural technique to cover timescales similar to those of electrophysiology measurements. Here, we used HS-AFM to directly monitor conformational dynamics of loop-6 in membrane-reconstituted OmpG at sub-millisecond temporal resolution. Then, we correlated these dynamics with single-channel recordings that provide kinetic information about channel gating. Furthermore, we performed molecular dynamics (MD) simulations to analyze pH-dependent gating dynamics and construct free-energy landscapes of OmpG. This multi-pronged analysis allowed us to establish the structure-dynamics-function relationship of a simple membrane protein. We hope that these technical advancements would open new avenues in structural biology community.