TY - JOUR
T1 - Purification of a membrane protein with conjugated engineered micelles
AU - Patchornik, Guy
AU - Danino, Dganit
AU - Kesselman, Ellina
AU - Wachtel, Ellen
AU - Friedman, Noga
AU - Sheves, Mordechai
N1 - Israel Science Foundation; Russell Berrie Nanotechnology Institute, TechnionWe would like to thank the Kimmelman Center for Biomolecular Structure and Assembly at the Weizmann Institute of Science for its generous support (to M.S.). M.S. holds the Katzir-Makineni chair in chemistry. D.D. thanks the Israel Science Foundation and the Russell Berrie Nanotechnology Institute, Technion for their support. We thank Dr. Shira Albeck for providing us the E. coli lysate.
PY - 2013/7/17
Y1 - 2013/7/17
N2 - A novel method for purifying membrane proteins is presented. The approach makes use of engineered micelles composed of a nonionic detergent, β-octylglucoside, and a hydrophobic metal chelator, bathophenanthroline. Via the chelators, the micelles are specifically conjugated, i.e., tethered, in the presence of Fe2+ ions, thereby forming micellar aggregates which provide the environment for separation of lipid-soluble membrane proteins from water-soluble proteins. The micellar aggregates (here imaged by cryo-transmission electron microscopy) successfully purify the light driven proton pump, bacteriorhodopsin (bR), from E. coli lysate. Purification takes place within 15 min and can be performed both at room temperature and at 4 C. More than 94% of the water-soluble macromolecules in the lysate are excluded, with recovery yields of the membrane protein ranging between 74% and 85%. Since this approach does not require precipitants, high concentrations of detergent to induce micellar aggregates, high temperature, or changes in pH, it is suggested that it may be applied to the purification of a wide variety of membrane proteins.
AB - A novel method for purifying membrane proteins is presented. The approach makes use of engineered micelles composed of a nonionic detergent, β-octylglucoside, and a hydrophobic metal chelator, bathophenanthroline. Via the chelators, the micelles are specifically conjugated, i.e., tethered, in the presence of Fe2+ ions, thereby forming micellar aggregates which provide the environment for separation of lipid-soluble membrane proteins from water-soluble proteins. The micellar aggregates (here imaged by cryo-transmission electron microscopy) successfully purify the light driven proton pump, bacteriorhodopsin (bR), from E. coli lysate. Purification takes place within 15 min and can be performed both at room temperature and at 4 C. More than 94% of the water-soluble macromolecules in the lysate are excluded, with recovery yields of the membrane protein ranging between 74% and 85%. Since this approach does not require precipitants, high concentrations of detergent to induce micellar aggregates, high temperature, or changes in pH, it is suggested that it may be applied to the purification of a wide variety of membrane proteins.
UR - http://www.scopus.com/inward/record.url?scp=84880394343&partnerID=8YFLogxK
U2 - 10.1021/bc400069w
DO - 10.1021/bc400069w
M3 - مقالة
C2 - 23758098
SN - 1043-1802
VL - 24
SP - 1270
EP - 1275
JO - Bioconjugate Chemistry
JF - Bioconjugate Chemistry
IS - 7
ER -