Seminar series of the Bionanoscience (BN) department of Delft University of Technology, including lunch.

This week's speaker: Christos Gogou on "Toward resolving protein voltage-sensitivity using cryo-EM". 

Abstract: Action potentials are a vital characteristic of neurons for proper brain function. Mid-20th century electrophysiology studies have early on demonstrated that ion channels open and close during such dynamic action potentials. Since then, many more (neuronal) transmembrane proteins were found to be voltage-sensitive. Although it is known that such proteins alter their structural conformations, and hence their cellular function, along changes in membrane potential, the underlying structural mechanisms remain poorly understood. Here, I aim to make use of the immense advances in protein structure determination through electron cryo-microscopy (cryo-EM) to reconstruct voltage-sensitive proteins in the presence of an actual membrane voltage. I will present the first steps toward a general cryo-EM-compatible method for capturing voltage-dependent conformations of membrane-inserted proteins. The lipid bi-layers of liposomes are deployed as cellular membrane-mimetic system with controllable size and chemical content. Manipulation of intra- and extra-liposomal ionic concentrations, and selective K+-permeation then allow tuning of the membrane potential. As a proof-of-principle, a fluorescent voltage indicator called ASAP2s was selected, successfully purified, and inserted into these liposomes. Functional assays to test whether purified ASAP2s maintains its voltage-sensitivity are ongoing.  A general method to apply membrane potentials for structural studies further unlocks the potential of cryo-EM, and may generate new insights in the function of (neuronal) transmembrane proteins in health and disease.