PloS one, 2021-02, Vol.16 (2), p.e0247689-e0247689
2021
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Details
- Autor(en) / Beteiligte
- Titel
- Refolding and characterization of two G protein-coupled receptors purified from E. coli inclusion bodies
- Ist Teil von
- PloS one, 2021-02, Vol.16 (2), p.e0247689-e0247689
- Ort / Verlag
- United States: Public Library of Science
- Erscheinungsjahr
- 2021
- Quelle
- Free E-Journal (出版社公開部分のみ）
- Beschreibungen/Notizen
- Aiming at streamlining GPCR production from E. coli inclusion bodies for structural analysis, we present a generic approach to assess and optimize refolding yield through thermostability analysis. Since commonly used hydrophobic dyes cannot be applied as probes for membrane protein unfolding, we adapted a technique based on reacting cysteins exposed upon thermal denaturation with fluorescent 7-Diethylamino-3-(4-maleimidophenyl)-4-methylcoumarin (CPM). Successful expression, purification and refolding is shown for two G protein-coupled receptors (GPCR), the sphingosine-1-phosphate receptor S1P1, and the orphan receptor GPR3. Refolded receptors were subjected to lipidic cubic phase crystallization screening.
- Sprache
- Englisch
- Identifikatoren
- ISSN: 1932-6203eISSN: 1932-6203DOI: 10.1371/journal.pone.0247689Titel-ID: cdi_plos_journals_2492989979
- Format
- –
- Schlagworte
- Addictive behaviors, Bacterial proteins, Biology and Life Sciences, Biotechnology, Cell receptors, Centrifugation, Chromatography, Crystallization, Developmental biology, E coli, Editing, Escherichia coli, Fluorescence, Freeze thaw cycles, Freeze-thawing, G protein-coupled receptors, Glucose, Glycerol, Hydrophobicity, Inclusion bodies, Kinases, Ligands, Lysozyme, Membrane proteins, Physical Sciences, Physiological aspects, Potassium chloride, Potassium phosphate, Potassium phosphates, Protein expression, Protein folding, Protein research, Proteins, R&D, Receptors, Research & development, Reviews, Sodium chloride, Sodium hydrogen phosphate, Sphingosine 1-phosphate, Streamlining, Structural analysis, Structure, Thermal denaturation, Thermal stability