المؤلفون: |
Stuebler M; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States.; University of Natural Resources and Life Sciences, Vienna 1180, Austria., Manzer ZA; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States., Liu HY; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States., Miller J; School of Integrative Plant Science, Cornell University, Ithaca, New York 14853, United States., Richter A; Boyce Thompson Institute, Ithaca, New York 14853, United States., Krishnan S; Boyce Thompson Institute, Ithaca, New York 14853, United States., Selivanovitch E; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States., Banuna B; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States., Jander G; Boyce Thompson Institute, Ithaca, New York 14853, United States., Reimhult E; University of Natural Resources and Life Sciences, Vienna 1180, Austria., Zipfel WR; Meinig School of Biomedical Engineering, Cornell University, Ithaca, New York 14853, United States., Roeder AHK; School of Integrative Plant Science, Cornell University, Ithaca, New York 14853, United States.; Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, New York 14853, United States., Piñeros MA; School of Integrative Plant Science, Cornell University, Ithaca, New York 14853, United States.; Robert W. Holley Center for Agriculture & Health, ARS-USDA, Ithaca, New York 14853, United States., Daniel S; RF Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States. |
مستخلص: |
The cell plasma membrane is a two-dimensional, fluid mosaic material composed of lipids and proteins that create a semipermeable barrier defining the cell from its environment. Compared with soluble proteins, the methodologies for the structural and functional characterization of membrane proteins are challenging. An emerging tool for studies of membrane proteins in mammalian systems is a "plasma membrane on a chip," also known as a supported lipid bilayer. Here, we create the "plant-membrane-on-a-chip,″ a supported bilayer made from the plant plasma membranes of Arabidopsis thaliana , Nicotiana benthamiana , or Zea mays . Membrane vesicles from protoplasts containing transgenic membrane proteins and their native lipids were incorporated into supported membranes in a defined orientation. Membrane vesicles fuse and orient systematically, where the cytoplasmic side of the membrane proteins faces the chip surface and constituents maintain mobility within the membrane plane. We use plant-membrane-on-a-chip to perform fluorescent imaging to examine protein-protein interactions and determine the protein subunit stoichiometry of FLOTILLINs. We report here that like the mammalian FLOTILLINs, FLOTILLINs expressed in Arabidopsis form a tetrameric complex in the plasma membrane. This plant-membrane-on-a-chip approach opens avenues to studies of membrane properties of plants, transport phenomena, biophysical processes, and protein-protein and protein-lipid interactions in a convenient, cell-free platform. |