Functional and Structural Information of TRPV1

The research aims to gain structural and functional information about membrane proteins where the focus is currently on an ion channel called TRPV1. In humans, TRPV1 is responsible for pain sensation at elevated temperatures, during inflammation and after flesh wounds but many have also experienced the effect of this ion channel after eating chili peppers. Capsaicin, the pungent compound in chili have the ability to activate TRPV1 leading to a hot, slightly painful sensation when eating chili.

TRPV1 has been the subject for intense research and gained a large interest from the drug industry due to its involvement in pain regulation and as a possible drug target for treatment of chronic pain. A future drug candidate should block its response to inflammatory compounds but TRPV1 should still be activated at elevated temperatures since that is an important regulatory mechanis. In order to develop such a drug, we need to gain more knowledge about activation mechanism and ligand binding sites for this ion channel.

The activity of TRPV1 is studied with an electrophysiological technique called patch-clamp where currents are measured through the ion channel at different conditions. The current focus is to evaluate behavioral differences for TRPV1 due to changes in cellular membrane compositions which could have an effect on the level of activity. We have concluded that a decrease in cholesterol levels in the plasma membrane interferes with expansion of the pore opening of TRPV1, which is a phenomenon that occurs during prolonged activation of the ion channel.

In addition to functional studies, proteomic studies are also performed in order to find potential binding sites. By performing proteolysis of TRPV1 at short time scale and analyzing the digested peptides with mass spectrometry you could map exposed regions of the protein which often correlates with binding sites, giving clues to which regions are important for activation and regulation. Proteolysis is also a simple way to digest parts of the protein and can be further used when trying to evaluate the effect of different regions on ion channel activity. Here we perform tryptic digestions within a microfluidic flow cell followed by functional studies with fluorescence. The same proteolysis can also be performed during patch-clamp in order to correlate structure with detailed functional data.