The styrene-maleic acid copolymer: a versatile tool in membrane research

Abstract

A new and promising tool in membrane research is the detergent-free solubilization of membrane proteins by styrene-maleic acid copolymers (SMAs). These amphipathic molecules are able to solubilize lipid bilayers in the form of nanodiscs that are bounded by the polymer. Thus, membrane proteins can be directly extracted from cells in a water-soluble form while conserving a patch of native membrane around them. In this review article, we briefly discuss current methods of membrane protein solubilization and stabilization. We then zoom in on SMAs, describe their physico-chemical properties, and discuss their membrane-solubilizing effect. This is followed by an overview of studies in which SMA has been used to isolate and investigate membrane proteins. Finally, potential future applications of the methodology are discussed for structural and functional studies on membrane proteins in a near-native environment and for characterizing protein-lipid and protein-protein interactions.

Keywords: Lipid–protein interactions; Lipodisq; Membrane proteins; Native nanodiscs; SMALPs; Styrene–maleic acid copolymers.

Fig. 1

Membrane-mimetic systems for membrane protein stabilization. The protein is indicated in blue and lipids in bilayers are indicated in green. a Protein in detergent (red) micelle. b Protein stabilized by amphipol (orange). c Protein in bicelle (detergent in red). d Protein in nanodisc stabilized by MSP (purple). e Protein in nanodisc stabilized by SMA (yellow)

Fig. 2

Schematic representation of the synthesis of styrene–maleic anhydride (Reaction 1) and the preparation of styrene–maleic acid (Reaction 2) as illustrated here for a 1:1 styrene-to-maleic anhydride/acid molar ratio. When styrene is present in excess, the monomer sequence distribution in the polymer becomes more complex (see text for details)

Fig. 3

Dimensions of styrene–maleic acid/lipid particles (SMALPs) consisting of DMPC lipids and a SMA copolymer with a styrene–maleic acid ratio of 2, as determined from small-angle neutron scattering experiments (figure adapted from Jamshad et al. 2015b)

Fig. 4

Schematic representation of a turbidimetry experiment. Suspensions of large lipid vesicles show a high degree of light scattering and appear milky. The addition of SMA leads to a clearing of the suspension due the formation of smaller nanodiscs. The concomitant rapid decrease in light scattering can be followed by measuring the apparent absorbance of the sample. For model membranes containing lipids in a fluid phase, this process typically occurs on time scales of minutes or tens of minutes (Scheidelaar et al. 2015)

Fig. 5

Three-step model for the solubilization of lipid membranes by SMA copolymers. Initially, SMA binds to the surface of the membrane (I), which is modulated by SMA and salt concentration and the presence of negatively charged lipids (PX⁻). The next step consists of the insertion of the polymer molecules into the hydrophobic core of the membrane (II), driven by the hydrophobic effect. This process is modulated by the lipid packing and bilayer thickness. Finally, the membrane is solubilized and nanodiscs are formed (III). The kinetics of the solubilization are determined mainly by the second step and after SMA has penetrated into the hydrophobic core of the bilayer, the formation of nanodiscs is a downhill process (see text for details, figure adapted from Scheidelaar et al. 2015)

Fig. 6

Extraction of membrane proteins with native lipid environment by SMA. SMA additions leads to the formation of native nanodiscs containing different MPs or only lipid material. Subsequent affinity purification allows for the isolation of native nanodiscs with the protein of interest

Fig. 7

Different applications of native nanodiscs. Dark green arrows indicate conservation of the native environment, light green arrows display possibilities for the transfer to controlled environments. Red arrows display approaches that generally use synthetic environments. The dashed arrows represent approaches in which complexes of MPs with specific lipids (red) or other proteins (yellow) can be isolated from native (2) or synthetic (6) environments