Elucidating the mechanism of bicelles spreading to fabricate sup-ported membranes containing an active transmembrane protein

Abstract

Transmembrane proteins participate in numerous cellular processes, but their investigation is complicated by the compositional complexity of biological membranes. In particular, G protein-coupled receptors, which represent targets for nearly 50% of pharmaceuticals, require well-defined model membrane systems for functional studies and drug development. Bicelles are attractive assemblies, because they can host transmembrane proteins and spread into planar supported lipid bilayers on solid surfaces. We fabricated bicelles with rhod(opsin) (dark-adapted and photoactivated) from outer segments of rod photoreceptor cells and developed a new methodology for the modification and activation of a monolayer deposited on the gold surface to anchor the N-terminus of rhod(opsin) and to form membranes with a uniformly oriented membrane protein. Bicelle spreading into a floating membrane was monitored by surface enhanced infrared absorption spectroscopy and quartz crystal microbalance measurements. The IR spectra revealed characteristic lipid and protein absorption bands, confirming a native fold of rhod(opsin). However, structural parameters such as the membrane thickness, hydration, protein distribution, and the architecture of the spacer layer, could not be elucidated from our previous experiments. Neutron reflectometry offers a powerful approach to determine these parameters and to elucidate the compositional and structural changes during the formation of model membranes.