title: Purification of a membrane protein with conjugated engineered micelles.
abstract: 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 Fe(2+) 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.
authors.affiliation: Department of Biological Chemistry, Ariel University, 70400, Israel. guyp@ariel.ac.il
authors.forename: Guy
authors.initials: G
authors.lastname: Patchornik
authors.forename: Dganit
authors.initials: D
authors.lastname: Danino
authors.forename: Ellina
authors.initials: E
authors.lastname: Kesselman
authors.forename: Ellen
authors.initials: E
authors.lastname: Wachtel
authors.forename: Noga
authors.initials: N
authors.lastname: Friedman
authors.forename: Mordechai
authors.initials: M
authors.lastname: Sheves
mesh_terms: D001436:Bacteriorhodopsins / Q000302:isolation & purification; D008565:Membrane Proteins / Q000302:isolation & purification*; D008823:Micelles; D046529:Microscopy, Electron, Transmission
chemical_list: D008565:Membrane Proteins; D008823:Micelles; D001436:Bacteriorhodopsins