Membrane proteins: structure - function - methods of analysis

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Exploiting the power of the native nanodisc Membrane proteins are found in all biological membranes. They have many essential functions and their surface location make them ideal drug targets in pharmaceutical and biotechnology industries. Despite their great importance, scientists still struggle to study these macromolecules in a functionally active form that is compatible with experimentation. The manipulation of membrane proteins in vitro traditionally requires the use of detergents that fail to mimic the properties of the original biological membrane. Even the mildest ones lead to protein destabilization or loss of protein activity, in addition to destroying protein-protein associations. The styrene-maleic acid (SMA) lipid particle, also termed native- or SMA-nanodisc, is a new tool to extract proteins from membrane with minimal amount of disruption. During SMA-nanodisc generation, the styrene moieties intercalate between the lipid acyl chains to form a “bracelet” which encircles a small patch of lipid bilayer (~10-15 nm diameter). In this manner, membrane proteins and also protein complexes, together with their native lipids, can be extracted and purified from the native membrane. In addition to the inherent advantage of conserving the native lipid environment and protein-protein interactions, a wide variety of applications can be developed because these particles are stable and water-soluble. The native nanodisc is an exciting new development in the field of membrane biology. In this proposal, we will exploit the power of native nanodiscs in three different ways. We will 1) employ a proteomic-based method to identify and isolate novel membrane protein complexes that are unstable in detergent, 2) construct a library of SMA-nanodiscs to identify membrane receptor(s) of virtually any given ligand, and 3) develop a protocol to allow rapid identification of specific lipids surrounding membrane proteins of any type of cell. Each of these applications will contribute important new information to key cellular process that occurs at the membrane interface. Furthermore, since membrane proteins are becoming easier to obtain in their native state, our work might stimulate interest from the pharmaceutical sector, especially for membrane protein targets previously considered too difficult to manufacture efficiently for downstream applications such as drug discovery.