The Journal of General Physiology, Vol 75, 251-270, Copyright © 1980 by The Rockefeller University Press
Fusion of phospholipid vesicles with planar phospholipid bilayer membranes. II. Incorporation of a vesicular membrane marker into the planar membrane
FS Cohen, J Zimmerberg and A Finkelstein
Fusion of multilamellar phospholipid vesicles with planar phospholipid
bilayer membranes was monitored by the rate of appearance in the planar
membrane of an intrinsic membrane protein present in the vesicle membranes.
An essential requirement for fusion is an osmotic gradient across the
planar membrane, with the cis side (the side containing the vesicles)
hyperosmotic to the opposite (trans) side; for substantial fusion rates,
divalent cation must also be present on the cis side. Thus, the low fusion
rates obtained with 100 mM excess glucose in the cis compartment are
enhanced orders of magnitude by the addition of 5- 10 mM CaCl2 to the cis
compartment. Conversely, the rapid fusion rates induced by 40 mM CaCl2 in
the cis compartment are completely suppressed when the osmotic gradient
(created by the 40 mM CaCl2) is abolished by addition of an equivalent
amount of either CaCl2, NaCl, urea, or glucose to the trans compartment. We
propose that fusion occurs by the osmotic swelling of vesicles in contact
with the planar membrane, with subsequent rupture of the vesicular and
planar membranes in the region of contact. Divalent cations catalyze this
process by increasing the frequency and duration of vesicle-planar membrane
contact. We argue that essentially this same osmotic mechanism drives
biological fusion processes, such as exocytosis. Our fusion procedure
provides a general method for incorporating and reconstituting transport
proteins into planar phospholipid bilayer membranes.
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