Amphipols (APols) are short amphipathic polymers that can substitute for detergents to keep integral membrane proteins (MPs) water soluble. In this review, we discuss their structure and solution behavior; the way they associate with MPs; and the structure, dynamics, and solution properties of the resulting complexes. All MPs tested to date form water-soluble complexes with APols, and their biochemical stability is in general greatly improved compared with MPs in detergent solutions. The functionality and ligand-binding properties of APol-trapped MPs are reviewed, and the mechanisms by which APols stabilize MPs are discussed. Applications of APols include MP folding and cell-free synthesis, structural studies by NMR, electron microscopy and X-ray diffraction, APol-mediated immobilization of MPs onto solid supports, proteomics, delivery of MPs to preexisting membranes, and vaccine formulation.

One of the major obstacles to membrane protein (MP) structural studies is the destabilizing effect of detergents. Amphipols (APols) are short amphipathic polymers that can substitute for detergents to keep MPs water-soluble under mild conditions. In the present work, we have explored the feasibility of studying the structure of APol-complexed MPs by NMR. As a test MP, we chose the 171-residue transmembrane domain of outer MP A from Escherichia coli (tOmpA), whose x-ray and NMR structures in detergent are known. 2 H, 15 N-labeled tOmpA was produced as inclusion bodies, refolded in detergent solution, trapped with APol A8-35, and the detergent removed by adsorption onto polystyrene beads. The resolution of transverse relaxation-optimized spectroscopy-heteronuclear single-quantum correlation spectra of tOmpA͞A8-35 complexes was found to be close to that of the best spectra obtained in detergent solutions. The dispersion of chemical shifts indicated that the protein had regained its native fold and retained it during the exchange of surfactants. MP-APol interactions were mapped by substituting hydrogenated for deuterated A8-35. The resulting dipolar broadening of amide proton linewidths was found to be limited to the ␤-barrel region of tOmpA, indicating that A8-35 binds specifically to the hydrophobic transmembrane surface of the protein. The potential of this approach to MP studies by solution NMR is discussed. membrane proteins ͉ amphipols ͉ OmpA ͉ surfactant I ntegral membrane proteins (MPs) are involved in such essential cell functions as energy transduction, import and export of nutrients and drugs, signal detection, cell-to-cell communication, etc. They comprise 20-30% of the proteins encoded in the genome of cells and a majority of the targets of currently marketed drugs (1). A detailed knowledge of their structure is essential to understanding their function and dysfunction, as well as to a wide range of biomedical and biotechnological applications. The scarcity of high-resolution MP structures (which represent Ͻ0.3% of currently available structures) can be traced to three main factors: low levels of natural abundance, difficult overexpression, and a poor stability in the presence of detergent. Detergents are generally used to handle MPs in aqueous solutions, because the highly hydrophobic character of their transmembrane surface renders MPs water-insoluble. By adsorbing onto this surface, detergents make it hydrophilic (2). However, the dissociating character of detergents, combined with the need to maintain an excess of them, frequently results in more or less rapid inactivation of solubilized MPs (3).Inactivation by detergents is a particularly serious problem in the field of solution-state NMR for the following reasons: (i) to keep highly concentrated MPs (in the mM range) from aggregating, high concentrations of detergents must generally be used (usually in the 200-to 600-mM range; see, for instance, ref. 4); (ii) high temperatures are usually resorted to, to improve the resolution of the spectra; and ...