Conformational Chaperones for Structural Studies of Membrane Proteins Using Antibody Phage Display with Nanodiscs
Published in Structure. 2015 Dec 28. pii: S0969-2126(15)00503-1. [Epub ahead of print]. PMID: 26749445. PMCID: 4740257. Link to Pubmed page.
Core Facility: Synthetic Antigen Binder (SAB) Generation and Crystallography and Protein Production Core.
A major challenge in membrane biophysics is to define the mechanistic linkages between a protein’s conformational transitions and its function. We describe a novel approach to stabilize transient functional states of membrane proteins in native-like lipid environments allowing for their structural and biochemical characterization. This is accomplished by combining the power of antibody Fab-based phage display selection with the benefits of embedding membrane protein targets in lipid-filled nanodiscs. In addition to providing a stabilizing lipid environment, nanodiscs afford significant technical advantages over detergent-based formats. This enables the production of a rich pool of high-performance Fab binders that can be used as crystallization chaperones, as fiducial markers for single-particle cryoelectron microscopy, and as probes of different conformational states. Moreover, nanodisc-generated Fabs can be used to identify detergents that best mimic native membrane environments for use in biophysical studies.
- Nanodiscs allow for generation of multiple antibody fragments to membrane proteins
- Improved phage display protocol is fast, efficient, and modular
- Resulting antibodies probe protein conformations present in lipid environment
- Expands the toolbox for structural and functional studies of membrane proteins