Strategies for the cell-free expression of membrane proteins.
Cell-free expression offers an interesting alternative method to produce membrane proteins in high amounts. Elimination of toxicity problems, reduced proteolytic degradation and a nearly unrestricted option to supply potentially beneficial compounds like cofactors, ligands or chaperones into the reaction are general advantages of cell-free expression systems. Furthermore, the membrane proteins may be translated directly into appropriate hydrophobic and membrane-mimetic surrogates, which might offer significant benefits for the functional folding of the synthesized proteins. Cell-free expression is a rapidly developing and highly versatile technique and several systems of both, prokaryotic and eukaryotic origins, have been established. We provide protocols for the cell-free expression of membrane proteins in different modes including their expression as precipitate as well as their direct synthesis into detergent micelles or lipid bilayers.
Reckel
Sina
S
Centre for Biomolecular Magnetic Resonance, Institute for Biophysical Chemistry, University of Frankfurt/Main, Frankfurt/Main, Germany.
Sobhanifar
Solmaz
S
Durst
Florian
F
Löhr
Frank
F
Shirokov
Vladimir A
VA
Dötsch
Volker
V
Bernhard
Frank
F
D000818:Animals; D002474:Cell-Free System; D003902:Detergents / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008563:Membrane Lipids / Q000737:chemistry; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D014176:Protein Biosynthesis; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D017510:Protein Folding; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis; D014158:Transcription, Genetic
D003902:Detergents; D008563:Membrane Lipids; D008565:Membrane Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins