ArticleProtein science : a publication of the Protein Society2009
Production of functional bacteriorhodopsin by an Escherichia coli cell-free protein synthesis system supplemented with steroid detergent and lipid.
Article in Protein science : a publication of the Protein Society, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 21 papers.
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21 citing papers in PubMed, 62 citations in OpenAlex.
- Cell-Free Gene Expression: Methods and Applications.Chemical reviews · 2025Review
- Cyanorhodopsin-II represents a yellow-absorbing proton-pumping rhodopsin clade within cyanobacteria.The ISME journal · 2024Article
- Crystallographic insights into lipid-membrane protein interactions in microbial rhodopsins.Frontiers in molecular biosciences · 2024Article
- Review
- Cell-free electrophysiology of human VDACs incorporated into nanodiscs: An improved method.Biophysical reports · 2021Article
- Preparation of cationic proteoliposomes using cell-free membrane protein synthesis: the chaperoning effect of cationic liposomes.RSC advances · 2020Article
- Review
- Functional G-Protein-Coupled Receptor (GPCR) Synthesis: The Pharmacological Analysis of Human Histamine H1 Receptor (HRH1) Synthesized by a Wheat Germ Cell-Free Protein Synthesis System Combined with Asolectin Glycerosomes.Frontiers in pharmacology · 2018Article
- Structural Mechanism for Light-driven Transport by a New Type of Chloride Ion Pump, Nonlabens marinus Rhodopsin-3.The Journal of biological chemistry · 2016Article
- Cell-free methods to produce structurally intact mammalian membrane proteins.Scientific reports · 2016Article
- Comprehensive study of liposome-assisted synthesis of membrane proteins using a reconstituted cell-free translation system.Scientific reports · 2015Article
- Allosteric regulation of γ-secretase activity by a phenylimidazole-type γ-secretase modulator.Proceedings of the National Academy of Sciences of the United States of America · 2014Article
- Cell-free synthetic biology: thinking outside the cell.Metabolic engineering · 2012Review
- Optimization of purification and refolding of the human chemokine receptor CXCR1 improves the stability of proteoliposomes for structure determination.Biochimica et biophysica acta · 2012Article
- Toward rational design of protein detergent complexes: determinants of mixed micelles that are critical for the in vitro stabilization of a G-protein coupled receptor.Biophysical journal · 2011Article
- Robotic large-scale application of wheat cell-free translation to structural studies including membrane proteins.New biotechnology · 2011Article
- Production and partial purification of membrane proteins using a liposome-supplemented wheat cell-free translation system.BMC biotechnology · 2011Article
- Functional expression of a two-transmembrane HtrII protein using cell-free synthesis.Biophysics (Nagoya-shi, Japan) · 2011Article
- An economical method for producing stable-isotope labeled proteins by the E. coli cell-free system.Journal of biomolecular NMR · 2010Article
- Nonmicellar systems for solution NMR spectroscopy of membrane proteins.Current opinion in structural biology · 2010Review
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7 authors at 2 institutions in 1 country.
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No grant is acknowledged in the PubMed record.
Abstract
Cell-free expression has become a highly promising tool for the efficient production of membrane proteins. In this study, we used a dialysis-based Escherichia coli cell-free system for the production of a membrane protein actively integrated into liposomes. The membrane protein was the light-driven proton pump bacteriorhodopsin, consisting of seven transmembrane alpha-helices. The cell-free expression system in the dialysis mode was supplemented with a combination of a detergent and a natural lipid, phosphatidylcholine from egg yolk, in only the reaction mixture. By examining a variety of detergents, we found that the combination of a steroid detergent (digitonin, cholate, or CHAPS) and egg phosphatidylcholine yielded a large amount (0.3-0.7 mg/mL reaction mixture) of the fully functional bacteriorhodopsin. We also analyzed the process of functional expression in our system. The synthesized polypeptide was well protected from aggregation by the detergent-lipid mixed micelles and/or lipid disks, and was integrated into liposomes upon detergent removal by dialysis. This approach might be useful for the high yield production of functional membrane proteins.
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