Membrane protein expression in cell-free systems.
Cell-free expression has emerged as a promising tool for the fast and efficient production of membrane proteins. The rapidly growing number of successfully produced targets in combination with the continuous development of new applications significantly promotes the distribution of this technology. Membrane protein synthesis by cell-free expression does not appear to be restricted by origin, size or topology of the target, and its global application is therefore a highly valuable characteristic. The technology is relatively fast to establish in standard biochemical labs, and it does not require expensive equipment. Moreover, it enables the production of membrane proteins in completely new modes, like the direct translation into detergent micelles, which is not possible with any other expression system. In this protocol, we focus on the currently most efficient cell-free expression system for membrane proteins based on Escherichia coli extracts.
Schneider
Birgit
B
Centre for Biomolecular Magnetic Resonance, University of Frankfurt/Main, Institute for Biophysical Chemistry, Frankfurt/Main, Germany.
Junge
Friederike
F
Shirokov
Vladimir A
VA
Durst
Florian
F
Schwarz
Daniel
D
Dötsch
Volker
V
Bernhard
Frank
F
D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D012995:Solubility; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
D008565:Membrane Proteins; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases