A scalable, GFP-based pipeline for membrane protein overexpression screening and purification.
We describe a generic, GFP-based pipeline for membrane protein overexpression and purification in Escherichia coli. We exemplify the use of the pipeline by the identification and characterization of E. coli YedZ, a new, membrane-integral flavocytochrome. The approach is scalable and suitable for high-throughput applications. The GFP-based pipeline will facilitate the characterization of the E. coli membrane proteome and serves as an important reference for the characterization of other membrane proteomes.
Drew
David
D
Department of Biochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden.
Slotboom
Dirk-Jan
DJ
Friso
Giulia
G
Reda
Torsten
T
Genevaux
Pierre
P
Rapp
Mikaela
M
Meindl-Beinker
Nadja M
NM
Lambert
Wietske
W
Lerch
Mirjam
M
Daley
Daniel O
DO
Van Wijk
Klaas-Jan
KJ
Hirst
Judy
J
Kunji
Edmund
E
De Gier
Jan-Willem
JW
D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D049452:Green Fluorescent Proteins / Q000032:analysis* / Q000235:genetics; D013294:Lactococcus lactis / Q000235:genetics; D049408:Luminescent Agents / Q000032:analysis*; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification
D029968:Escherichia coli Proteins; D049408:Luminescent Agents; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins