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Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Monday Speaker Abstracts
12
Membrane Binding and Remodelling by the COPII Complex
Jan Auerswald, Sebastian Daum, Jan Ebenhan, Daniela Kruger, Mona Gross, Stefan Werner,
Kirsten Bacia
.
University of Halle, Halle, Germany.
COPII vesicles are responsible for transporting cargo from the ER towards the Golgi apparatus in
the secretory pathway. The small GTPase Sar1, which belongs to Sar1/Arf family of GTPases
and the Ras-superfamily, is an essential component in COPII-vesicle formation. Upon activation
with GTP, Sar1 binds to membranes, embedding an amphipathic helix into the proximal leaflet
of the bilayer. The exact role of GTP presence versus GTP hydrolysis in the formation in COPII
vesicle fission is still controversial. We study COPII vesicle formation in a bottom-up fashion on
S. cerevisiae
proteins using cryo-electron microscopy techniques, confocal imaging and
biophysical techniques. Cryo-EM shows strongly different COPII coat morphologies under GTP
hydrolyzing versus non-hydrolyzing conditions. More subtle differences are observed by cryo-
EM among reconstituted COPII samples under non-hydrolyzing conditions. Membrane binding
of the small GTPases Sar1 and Arf1 is studied using biophysical techniques. By combining
fluorescence correlation spectroscopy (FCS) with a Langmuir film balance, the protein’s
footprint in the proximal membrane leaflet is revealed. Dual-color fluorescence cross-correlation
spectroscopy analysis has been developed into a method for obtaining binding curves and testing
binding models of protein binding to freely diffusing vesicles. The higher specificity makes
fluorescence cross-correlation advantageous compared to previous fluorescence autocorrelation
analysis also for small vesicle applications.