Detergent-free ultrafast reconstitution of membrane proteins into lipid bilayers using fusogenic complementary-charged proteoliposomes

Journal article

Detergents are indispensable for delivery of membrane proteins into 30-100 nm small unilamellar vesicles, while more complex, larger model lipid bilayers are less compatible with detergents. Here we describe a strategy for bypassing this fundamental limitation using fusogenic oppositely charged liposomes bearing a membrane protein of interest. Fusion between such vesicles occurs within 5 min in a low ionic strength buffer. Positively charged fusogenic liposomes can be used as simple shuttle vectors for detergent-free delivery of membrane proteins into biomimetic target lipid bilayers, which are negatively charged. We also show how to reconstitute membrane proteins into fusogenic proteoliposomes with a fast 30-min protocol. Combining these two approaches, we demonstrate a fast assembly of an electron transport chain consisting of two membrane proteins from E. coli, a primary proton pump bo 3 -oxidase and F 1 F o ATP synthase, in membranes of vesicles of various sizes, ranging from 0.1 to > 10 microns, as well as ATP production by this chain.

Authors: Galkin, M; Russell, A; Vik, S; Berry, R; Ishmukhametov, R

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Journal of Visualized Experiments
• Journal: Journal of Visualized Experiments
• Volume: 134
• Article number: e56909
• Publication date: 2018-04-05
• DOI: 10.3791/56909
• ISSN: 1940-087X