Surfactant-free purification of membrane proteins with intact native membrane environment

Abstract

In order to study the structure and function of a protein, it is generally required that the protein in question is purified away from all others. For soluble proteins, this process is greatly aided by the lack of any restriction on the free and independent diffusion of individual protein particles in three dimensions. This is not the case for membrane proteins, as the membrane itself forms a continuum that joins the proteins within the membrane with one another. It is therefore essential that the membrane is disrupted in order to allow separation and hence purification of membrane proteins. In the present review, we examine recent advances in the methods employed to separate membrane proteins before purification. These approaches move away from solubilization methods based on the use of small surfactants, which have been shown to suffer from significant practical problems. Instead, the present review focuses on methods that stem from the field of nanotechnology and use a range of reagents that fragment the membrane into nanometre-scale particles containing the protein complete with the local membrane environment. In particular, we examine a method employing the amphipathic polymer poly(styrene-co-maleic acid), which is able to reversibly encapsulate the membrane protein in a 10 nm disc-like structure ideally suited to purification and further biochemical study.

Effect of Polymer Composition and pH on Membrane Solubilization by Styrene-Maleic Acid Copolymers

“…This has opened options to purify membrane proteins that are unstable in detergent micelles and to study native protein-lipid interactions by biochemical methods. A further advantage of these so-called native nanodisks is that they are small, with sizes in the range of 10-25 nm (4)(5)(6)(9)(10)(11)(12)(13)(14)(15). This makes them suitable to be characterized by a variety of biophysical approaches including UV/Vis-and fluorescence spectroscopy (5,6,8), as well as light scattering techniques (6,14).…”

Detergent-free purification of ABC (ATP-binding-cassette) transporters

“…Here we report the application of a recently developed system for detergent-free extraction of ABC transporters from their native membranes using a readily available, biocompatible and chemically stable polymer, polystyrene-co-maleic acid (SMA) [17][18][19]. The polymer inserts into the membrane isolating small discs of lipid bilayer that are encircled by the polymer, termed SMA lipid particles (SMALPs) [17].…”

The Styrene-Maleic Acid Copolymer: A Versatile Tool in Membrane Research

“…Such reconstitution is not straightforward however, especially because of the high affinity of SMA for lipids, which complicates their removal and may impair the formation of extended bilayers. It has been suggested that lowering the pH would solve this problem (Jamshad et al 2011). However, in our hands this approach was not successful (Koorengevel, Scheidelaar and Dörr, unpublished observations).…”