Recent progress in structure determination of trimeric ion channels and the sodium leak channel (Kschonsak et al., Nature, 2022; Yoder & Gouaux, eLife, 2020; McCarthy et al., Cell, 2019) provides an excellent framework to shed new light on these fascinating macromolecules. In particular, our lab employs a combination of electrophysiology, chemical biology, protein engineering and fluorescence spectroscopy to address crucial questions regarding the molecular function and pharmacology of these proteins:
- ElectrophysiologyWhen ions flow across a membrane they generate electrical currents. Although minute, they can be measured with great precision, down to the level of ions flowing through a single ion channel. In our lab we employ a variety of electrophysiological recording techniques, e.g. two-electrode voltage-clamp and patch-clamp methodologies, including single molecule and high throughput recordinds (Lynagh et al., eLife, 2017; Braun et al., PLoS Biol, 2021).
- Protein engineeringEmerging chemical biology techniques allow the site-directed incorporation of ncAAs or post-translational modifications via either non-sense suppression approaches (Braun et al, J Physiol, 2020) or protein semi-synthesis (Khoo & Galleano et al., Nat Comm, 2020, Galleano & Harms, PNAS, 2021). These modifications offer the ability to either incorporate subtle derivatives of naturally occurring amino acids or to confer entirely new properties to a given protein.
- FluorometryPatch-clamp fluorometry (PCF) and its cousin voltage-clamp fluorometry (VCF) allow labeling an ion channel with an organic dye. When the protein changes its conformation, the dye reports a change in environment by changing its brightness (Borg & Braun et al, PNAS, 2020; Heusser & Borg et al, eLife, 2022). We recently increased the sensitivity of this approach, allowing to track protein conformational changes independent of ionic currents with greater temporal resolution (Wulf & Pless, Cell Rep, 2018).