Abstract
Membrane proteins are involved in various cellular processes. However, purification of membrane proteins has long been a challenging task, as membrane protein stability in detergent is the bottleneck for purification and subsequent analyses. Therefore, the optimization of detergent conditions is critical for the preparation of membrane proteins. Here, we utilize analytical ultracentrifugation (AUC) to examine the effects of different detergents (OG, Triton X-100, DDM), detergent concentrations, and detergent supplementation on the behavior of membrane protein TmrA. Our results suggest that DDM is more suitable for the purification of TmrA compared with OG and TritonX-100; a high concentration of DDM yields a more homogeneous protein aggregation state; supplementing TmrA purified with a low DDM concentration with DDM maintains the protein homogeneity and aggregation state, and may serve as a practical and cost-effective strategy for membrane protein purification.
Highlights
Membrane proteins account for ~20–30% of eukaryotic proteome and play crucial roles in a wide variety of cellular functions, such as regulating transmembrane ion transport, sensing and transmitting chemical or electrical signals, mediating cellular attachment, and modulating membrane lipid composition
The results suggest that the choice of detergent and the concentration of detergent dictate the behavior of TmrA
Nonionic detergents are generally mild and nondenaturing, without affecting the native conformations of membrane proteins [30,31]. Due to their amphiphilic nature, these mild detergents can improve the solubility of membrane proteins
Summary
Membrane proteins account for ~20–30% of eukaryotic proteome and play crucial roles in a wide variety of cellular functions, such as regulating transmembrane ion transport, sensing and transmitting chemical or electrical signals, mediating cellular attachment, and modulating membrane lipid composition. They are classified into integral and peripheral membrane proteins. Integral membrane proteins are embedded in the lipid bilayer, while peripheral membrane proteins are weakly associated with the hydrophilic surfaces of the lipid bilayer [1,2]. In the process of membrane protein purification, detergents are used to solubilize membrane proteins and to shield their hydrophobic surfaces in aqueous solutions. It is critical to choose the appropriate type and concentration of detergent in the interest of cost and efficiency [6,7]
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