Quality control of overexpressed membrane proteins.
Overexpression of membrane proteins in Escherichia coli frequently leads to the formation of aggregates or inclusion bodies, which is undesirable for most studies. Ideally, one would like to optimize the expression conditions by monitoring simultaneously and rapidly both the amounts of properly folded and aggregated membrane protein, a requirement not met by any of the currently available methods. Here, we describe a simple gel-based approach with green fluorescent protein as folding indicator to detect well folded and aggregated proteins simultaneously. The method allows for rapid screening and, importantly, pinpointing the most likely bottlenecks in protein production.
Geertsma
Eric R
ER
Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute and Zernike Institute for Advanced Materials, University of Groningen, Nijenborgh 4, 9747 AG, Groningen, The Netherlands.
Groeneveld
Maarten
M
Slotboom
Dirk-Jan
DJ
Poolman
Bert
B
D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008565:Membrane Proteins / Q000235:genetics*; D017510:Protein Folding; D011786:Quality Control
D008565:Membrane Proteins; D049452:Green Fluorescent Proteins