title: Folding kinetics of an alpha helical membrane protein in phospholipid bilayer vesicles.
abstract: We report a detailed kinetic study of the folding of an alpha-helical membrane protein in a lipid bilayer environment. SDS denatured bacteriorhodopsin was folded directly into phosphatidylcholine lipid vesicles by stopped-flow mixing. The folding kinetics were monitored with millisecond time resolution by time-resolving changes in protein fluorescence as well as in the absorption of the retinal chromophore. The kinetics were similar to those previously reported for folding bacteriorhodopsin in detergent or lipid micelles, except for the presence of an additional apoprotein intermediate. We suggest this intermediate is a result of the greater internal two-dimensional pressure present in these lipid vesicles as compared to micelles. These results lay the groundwork for future studies aimed at understanding the mechanistic origin of the effect of lipid bilayer properties on protein folding. Furthermore, the use of biologically relevant phosphatidylcholine lipids, together with a straightforward rapid mixing process to initiate the folding reaction, means the method is generally applicable, and thus paves the way for an improved understanding of the in vitro folding of transmembrane alpha-helical proteins.
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK.
authors.forename: Samantha J
authors.initials: SJ
authors.lastname: Allen
authors.forename: A Rachael
authors.initials: AR
authors.lastname: Curran
authors.forename: Richard H
authors.initials: RH
authors.lastname: Templer
authors.forename: Wim
authors.initials: W
authors.lastname: Meijberg
authors.forename: Paula J
authors.initials: PJ
authors.lastname: Booth
mesh_terms: D007700:Kinetics; D008051:Lipid Bilayers; D008565:Membrane Proteins / Q000737:chemistry*; D010743:Phospholipids / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
chemical_list: D008051:Lipid Bilayers; D008565:Membrane Proteins; D010743:Phospholipids