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Expression,stabilization and purification of membrane proteins via diverse protein synthesis systems and detergents involving cell-free associated with self-assembly peptide surfactants
Authors:Xuan Zheng  Shuangshuang Dong  Jie Zheng  Duanhua Li  Feng Li  Zhongli Luo
Affiliation:1. College of Basic Medical Sciences, Molecular Medicine and Cancer Research Center, Chongqing Medical University, Chongqing, China;2. College of laboratory Medicine, Chongqing Medical University, Chongqing, China;3. Sichuan Industrial Institute of Antibiotics, Chengdu University, Chengdu, China;4. China Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou, China
Abstract:G-protein coupled receptors (GPCRs) are involved in regulating most of physiological actions and metabolism in the bodies, which have become most frequently addressed therapeutic targets for various disorders and diseases. Purified GPCR-based drug discoveries have become routine that approaches to structural study, novel biophysical and biochemical function analyses. However, several bottlenecks that GPCR-directed drugs need to conquer the problems including overexpression, solubilization, and purification as well as stabilization. The breakthroughs are to obtain efficient protein yield and stabilize their functional conformation which are both urgently requiring of effective protein synthesis system methods and optimal surfactants. Cell-free protein synthesis system is superior to the high yields and post-translation modifications, and early signs of self-assembly peptide detergents also emerged to superiority in purification of membrane proteins. We herein focus several predominant protein synthesis systems and surfactants involving the novel peptide detergents, and uncover the advantages of cell-free protein synthesis system with self-assembling peptide detergents in purification of functional GPCRs. This review is useful to further study in membrane proteins as well as the new drug exploration.
Keywords:ac-A6K-CONH2, AAAAAAK-CONH2   ConA, lectin concanavalin A   Dcytb, cytochrome b   DDM, n-dodecyl-β-d-maltoside   E. coli, Escherichia coli   FSEC-TS, fluorescence-detection size-exclusion chromatography-based thermostability assay   GPCRs, G-protein coupled receptors   hFPR3, human formyl peptide receptor   hPOMGnT1, human protein of O-linked mannose β-1,2-N-acetylglucosaminyltransferase   hTAAR5, human trace-amine receptor   LCP, lipid cubic phase   MBCD, methyl-beta-cyclodextrin   MBP, maltose-binding protein   NAPols, nonionic amphiphols   OG, n-octyl-ß  -d-gluco-pyranoside   ORs, olfactory receptors   PS-I, photosystem I   SDC, sample displacement chromatography   Sf21, Spodoptera frugiperda 21   TAAR, human trace amine-associated receptor   TF, trigger factor   T4L, T4 lysozyme   VNRs, human vomeronasal receptors
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