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Magic™ Cell-free Protein Expression in E. coli Lysate

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Creative Biolabs is a world-leading service provider of cell-free membrane protein expression system. We have developed multiple advanced cell-free systems for membrane protein expression including but not limited to:


Amongst them, E. coli expression system represents the most robust, efficient and low-cost option. Our experts now offer integrated service of Magic™ membrane protein expression in E. coli cell-free expression system for our worldwide clients.

In vitro translation systems derived from a wide variety of cellular origins have been described in many literatures and have been widely used in laboratories. Conventional in vivo membrane protein expression systems are often limited by host cell toxicity, low yield, and purification using detergents. Compared with conventional cell-based systems, the cell-free system is a novel method which removes the cell membrane to leave the cytosolic and organelle components. E. coli lysate represents one earliest and most classic prokaryotic for cell-free expression. This cell-free production system, needed for amino acid incorporation in the presence of soluble tRNA and ribosome, activates an E. coli-derived cell lysates for transcription and translation. Ideally, extracts prepared from E. coli cells can produce between half and one milligram per milliliter of a reporter protein in a test tube after a few hours of incubation. T3 and T7 bacteriophage RNA polymerases are often used to achieve high protein yield. The E. coli cell-free expression system has following advantages:


Schematic of phosphoprotein synthesis from E. coli lysate. Sep-OTS: genetically encoded phosphoserine; CFPS: cell-free protein synthesis; NTPs: nucleoside triphosphates; lysis: breaking down cell membrane Figure 1. Schematic of phosphoprotein synthesis from E. coli lysate. Sep-OTS: genetically encoded phosphoserine; CFPS: cell-free protein synthesis; NTPs: nucleoside triphosphates; lysis: breaking down cell membrane (Nature Communications, 2015)

Currently, the E. coli cell-free expression system has been applied for the production of a broad variety of membrane proteins with regard to size, function, origin and topology, for example, GPCRs, ion channels, transporters. Applications on the record include X-ray crystallography, NMR structural analysis, functional studies, and the characterization of large membrane protein assemblies. For X-ray (selenomethionine substitution) and NMR (stable-isotope labeling), higher labeling/substitution efficiency can be achieved by E. coli cell-free expression system compared with cell-based expression systems.

Creative Biolabs is highly experienced in membrane proteins expression based on our first-class cell-free expression platform. Our scientists are professional in tailoring custom-oriented plans to provide ultimate satisfaction in high-quality membrane protein products. Please feel free to inquiry us for more information.

References

  1. Henrich E, Hein C, Dötsch V, et al. (2015) Membrane protein production in Escherichia coli cell‐free lysates. FEBS letters, 589(15), 1713-1722.
  2. Alkhalfioui F, Logez C, Bornert O, Wagner R (2011) Production of Membrane Proteins – Strategies for Expression and Isolation (Robinson AS, ed), Wiley-VCH: 1
  3. Oza JP, Aerni HR, Pirman NL, et al. (2015) Robust production of recombinant phosphoproteins using cell-free protein synthesis. Nature communications, 6.



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