Engineered nanodiscs for structural studies of the interaction of venom peptides with voltage-gated ion channels in a near native environment

AZ Zhang, XJ Jia and MM Mobli

Center for Advanced Imaging, The University of Queensland

Voltage-gated ion channels are a superfamily of membrane proteins encoded by more than 143 genes in the human genome, making it one of the largest superfamilies of signal transduction proteins. These ion channels are recognised as among the most common and important, yet underutilised, drug targets. Unsurprisingly, there has been a surge of biophysical studies in the recent decades that aim to assess the interactions of these ion channels with potential inhibitors such as peptide toxins. Structural studies of membrane proteins are typically undertaken in detergent micelles. This model, however, presents uncertainties in its representation of the native phospholipid bilayer environment. Recently, soluble lipid bilayer platforms called nanodiscs (NDs) have been utilised to study membrane proteins in an attempt to achieve a more native environment. Nanodiscs consist of a membrane scaffold protein (MSP) wrapped around a lipid bilayer and embedded membrane protein, forming disc-like rafts that essentially mimic the flat phospholipid bilayer. NDs can stabilise membrane proteins in more native conformations, and can ultimately be used to derive more physiologically relevant functional and structural data. Here, we have used modified NDs appropriate for study by NMR spectroscopy to gain further insight into lipid binding and channel inhibition of the natural ligands of voltage-gated ion channels. The peptide-binding voltage-sensor domain (VSD) of a potassium channel, KvAP, was re-suspended in a nanodisc. We used NMR spectroscopy to study the interactions of a known peptide inhibitor, VSTx1, with the KvAP VSD. The study provides one of the first atomic resolution details of the binding of a peptide toxin to a VSD embedded in a lipid bilayer.