• Amphipol A8-35
• Protein Trapping
• Properties of MP/APol complexes
• Functionalized A8-35
• Others APols
• Applications
• Distribution/Training
• References

• Overview
• Folding and Immobilization of Membrane Proteins
• Applications to NMR studies
• Biomedical Applications
• Applications to microscopy

8_ Summary

The high quality of NMR signals observed with membrane proteins associated to amphipols opens perspectives of complete structural determination of these proteins, provided their size is compatible with solution NMR studies. Amphipols thus join bicelles (Raschle et al (2010), Vold et al (1996)) or nanodiscs (Bayburt et al (2002), Denisov et al (2004)) as workable alternatives to detergents. Membrane protein/amphipol complexes are slightly larger than the smallest membrane proteins/detergent complexes. However, the search for the fastest tumbling times for solution-state NMR studies, which was justified a decade ago, appears not so critical today, with the progress of NMR equipment and methodology, and with the help of improved isotopic labeling strategies (Tugarinov et al (2006)).

Depending on the MP under study, a slight loss in resolution might be more than compensated for by the improvement in stability, which allows longer collection times. Futhermore, a higher stability may also make it possible to improve the resolution by raising the temperature. Media milder than detergents, whether they provide a bilayer-like environment, like bicelles and nanodiscs, or favor the retention of membrane proteins-bound lipids, as amphipols seem to do, are likely to better preserve native-like structural features. This is suggested by differences in protein signal chemical shifts between membrane protein/detergent complexes and either membrane protein/bicelle complexes ( Chou et al (2002), Lee et al (2008)) or membrane protein/nanodisc complexes (Glück et al (2009), Raschle et al (2009)).

It will be interesting to examine how well amphipols fare in this respect. A notable advantage of amphipols is the simplicity of preparation and handling of membrane protein/amphipol complexes. Membrane protein/bicelle complexes indeed require a strict control of the ratio of long chain vs. short chain lipids or detergents all along the preparation and data collection to ensure the presence of small and monodisperse bicellar particles (Triba et al (2005), Sanders et al (2006)). This control is far from trivial when buffer exchange or protein concentration steps are required. Membrane protein/nanodisc complexes are simpler to handle once they have been formed, but their preparation involves the production and purification of substantial amounts of lipoproteins, especially if one aims to capture a single membrane proteins per nanodisc, and some purification work is required to obtain monodisperse preparations (see e.g. ref. Ritchie et al (2009)).

From a practical point of view, membrane protein complexes with A8-35, SAPols and NAPols afford comparable resolution. A marked advantage of A8-35 or SAPols is that they are relatively easy to deuterate, which is a great asset for certain NMR studies. They would have to be preferred to NAPols should it be necessary to work with unprotonated amphipols, because perdeuteration of glucosylated NAPols would be difficult and costly. On the other hand, existing data suggest that NAPols may be even milder towards membrane proteins than A-35 or SAPols (see ref.Bazzacco et al. (2012)).