Mempro™ Detergent-Free Pentameric Ligand-Gated Ion Channel Production

Creative Biostructure provides specific custom Mempro™ pentameric ligand-gated ion channel (pLGIC) production services using detergent-free expression system.

The pLGIC, a single neurotransmitter receptor superfamily, mediates fast chemical transmission by converting nerve signals into an electrical response. It has provent that pLGICs are a group of transmembrane ion channels that open to allow ions such as Na⁺, K⁺, Ca²⁺, or Cl⁻ to pass through the cell membrane, which are composed of five homologous subunits. And theses subunits consist of at least two different domains, including a transmembrane domain which forms the channel gate and the ion selective filter, and an extracellular domain which includes the ligand binding location (an allosteric binding site). In addition, pLGICs are divided into three classes: Cys-loop receptors, Ionotropic glutamate receptors and ATP-gated channels. Traditional approaches for pLGIC purification need extraction from the membrane into detergent micelles. However, detergent can induce conformational change on membrane proteins. Our alternative detergent-free expression system can best reserve the native conformation of pLGICs for further structural studies.

Figure 1. Ligand-gated ion channels can be divided into three large multigene families. (Nature Review Neuroscience, 2001)

Creative Biostructure has extensive experience in high-yield pLGIC production using detergent-free membrane protein expression system, we can employ various strategies for Mempro™ detergent-free transmember protein production, including:

• Mempro™ pLGIC production using nanodiscs

  Nanodiscs are the self-assembly system to stabilize membrane proteins removed from the membrane by membrane scaffold proteins (MSPs) in no need of detergent. Creative Biostructure can reconstitute pLGICs into nanodiscs in detergent-free process: Expressing the pLGICs in a cell-free system with the addition of pre-assembled nanodiscs.

• Mempro™ pLGIC production using amphipols

  The amphipathic amphipols have the ability to “trap” around transmembrane regions of proteins, allowing them to stay folded. Creative Biostructure can utilize A8-35 to solubilize pLGICs during purification step.

• Mempro™ pLGIC production using poly (styrene-co-maleic acid) lipid particles (SMALPs)

The SMALPs are self-assembled by the simple addition of the SMA co-polymers. At neutral or alkaline pH, a disc-like structure assembles itself, encapsulating pLGICs in a form amenable to be purified.

These novel detergent-free technologies for pLGIC production can be obtained easily, and enabling more comprehensively structural and functional studies.

Creative Biostructure provides other various Mempro™ membrane protein production services. Please feel free to contact us for a detailed quote.

References:
B. S. Khakh, et al. (2001). Molecular physiology of p2x receptors and atp signalling at synapses. Nature Review Neuroscience, 2: 165-174.
C. Tribet, et al. (1996). Amphipols: Polymers that keep membrane proteins soluble in aqueous solutions. Proc Natl Acad Sci U. S. A. 93: 15047-15050.
J. A. Dent (2010). The evolution of pentameric ligand-gated ion channels. Adv. Exp. Med. Biol., 683: 11-23.
M. Jamshad, et al. (2011). Surfactant-free purification of membrane proteins with intact native membrane environment. Biochem. Soc. Trans., 39: 813-818.