CLATHRIN-INDUCED PH-DEPENDENT FUSION OF PHOSPHATIDYLCHOLINE VESICLES

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  • 1 January 1983
    • journal article
    • research article
    • Vol. 258  (5) , 3409-3415
Abstract
Interaction of [rat liver] clathrin coat protein with dioleoylphosphatidylcholine (DOPC) vesicles at pH 6.5 and below results in the formation of stable vesicle-clathrin complexes. Gel chromatography and sedimentation analysis show that the interaction of clathrin coat protein with unilamellar DOPC vesicles at pH = 6.0 results in the formation of larger structures. EM and an increase in trapped volume of both sucrose and inulin show that those larger structures represent fused bilayers. The mixing of membrane lipid as a result of membrane fusion was examined using resonance energy transfer between 2 fluorescent lipid probes incorporated into the same vesicle membrane. At a protein:lipid ratio of 1:500 there was 50% vesicle-vesicle fusion, at pH 6.0, as indicated by the change in efficiency of energy transfer between the fluorescent probes. Fusion was completed within 60 s. A number of other proteins (ovalbumin, rabbit IgG, trypsin, pronase, calmodulin, tubulin, synexin, bovine serum albumin) at 10-fold or higher concentrations did not induce fusion of DOPC vesicles, either at pH 7.4 or at pH 6.0. This system provides a model for pH-dependent and protein-mediated fusion of uncharged lipid bilayers.