A giant liposome for single-molecule observation of conformational changes in membrane proteins
Journal article, 2009
We present an experimental system that allows visualization of conformational changes in membrane proteins at the single-molecule level. The target membrane protein is reconstituted in a giant liposome for independent control of the aqueous environments on the two sides of the membrane. For direct observation of conformational changes, an extra-liposomal site(s) of the target protein is bound to a glass surface, and a probe that is easily visible under a microscope, such as a micron-sized plastic bead, is attached to another site on the intra-liposomal side. A conformational change, or an angular motion in the tiny protein molecule, would manifest as a visible motion of the probe. The attachment of the protein on the glass surface also immobilizes the liposome, greatly facilitating its manipulation such as the probe injection. As a model system, we reconstituted ATP synthase (FOF1) in liposomes tens of mu m in size, attached the protein specifically to a glass surface, and demonstrated its ATP-driven rotation in the membrane through the motion of a submicron bead. (c) 2009 Elsevier B.V. All rights reserved.
escherichia-coli
c-subunit ring
Single-molecule
stepwise rotation
ATP synthase
Membrane protein
unilamellar vesicles
change
flagellar motor
kinetic mechanism
mitochondrial adenosine-triphosphatase
channel
Giant liposome
Conformational
atp synthase
resonance energy-transfer
Author
Y. Onoue
Waseda University
The Graduate University for Advanced Studies (SOKENDAI)
T. Suzuki
Tokyo Institute of Technology
Japan Science and Technology Agency
M. Davidson
Chalmers
Mattias Karlsson
Chalmers
Owe Orwar
Chalmers, Chemical and Biological Engineering, Physical Chemistry
M. Yoshida
Japan Science and Technology Agency
Tokyo Institute of Technology
K. Kinosita
Waseda University
Biochimica et Biophysica Acta - Biomembranes
0005-2736 (ISSN) 1879-2642 (eISSN)
Vol. 1788 6 1332-1340Subject Categories
Physical Chemistry
Chemical Sciences
DOI
10.1016/j.bbamem.2009.01.015