Lysozyme-induced fusion of liposomes with erythrocyte ghosts at acidic pH.

Abstract

Lysozyme that was covalently bound to the outer surface of sonicated vesicles induced fusion of the vesicles with human white erythrocyte ghosts. The kinetics of membrane mixing were evaluated by the resonance-energy-transfer method using L-a-dipalmitoyl phosphatidylethanolamine labeled at the free amino group with the energy donor 7-nitro-2,1,3-benzoxadiazol-4-yl or with the energy acceptor tetramethylrhodamine. The equilibrium state after fusion was characterized by using fluorescence photobleaching and recovery techniques. Rates and equilibrium percentages of fusion were maximal at the pH optimum of the enzyme, and rates were strongly reduced by the addition of N,N',N"-triacetylchitotriose, a competitive inhibitor of lysozyme. An apparent activation energy of 28 ± kcal/mol was obtained for the lipid-mixing process. At 370C, the fusion half-time was 0.5 min. After 30 min at 370C, 40% of the labeled lipids initially present in the fusion mixture had a lateral diffusion constant, D, of 1.1 ± 0.5 x 10-9 cm2.sec-1 in the ghost membrane. The strong induction of fusion at the lysozyme pH optimum was not observed in the absence of lysozyme or when free lysozyme was added to the solution. Bound lysozyme did not induce fusion of electrically neutral liposomes with each other. These observations indicate that it is the liposome-bound lysozyme that induces fusion between liposomes and erythrocyte ghosts.

Calcium-dependent fusion of the plasma membrane fraction from human neutrophils with liposomes

“…The apparent lack of Ca2+-dependent fusion of neutrophil granules with membranes suggests that additional conditions and/or cofactors may be required to promote Ca2÷-dependent fusion of granules with plasma membranes. This observation is consistent with recent findings that have shown that proteins, fatty acids, nucleotides and phosphorylation reactions may be required for membrane fusion and exocytosis [60][61][62][63][64][65]. Thus, determining the fusion sensitivity of isolated membrane components to Ca 2÷ should aid in further elucidation of the cofactor requirements of exocytic membrane fusion in the neutrophil.…”

Membrane penetration of Sendai virus glycoproteins during the early stages of fusion with liposomes as determined by hydrophobic photoaffinity labeling.

“…The very similar F/HN labeling ratios observed under various conditions of temperature and pH suggest a common fusion mechanism for both vesicle types. Furthermore, the higher HN labeling relative to F observed at low pH supports the hypothesis that HN mediates fusion at pH 5.0 by a low-pHinduced conformational change allowing hydrophobic interaction with the target membrane (14,28), and is consistent with the model for fusion mediated by water-soluble proteins at low pH (12,29,30).…”

Lysozyme interaction with negatively charged lipid bilayers: protein aggregation and membrane fusion

“…[4][5][6] HEWL is structurally homologous to human lysozyme, involved in several destabilizing mutations-related to protein deposition into amyloid plaques found in the liver, kidneys, and spleen of patients affected by non-neuropathic hereditary amyloidosis. [7][8][9][10][11] Because of its homology to human lysozyme, HEWL has been extensively used as a model in studies for a number of investigations: protein adsorption at interfaces, 12,13 membrane fusion, [14][15][16][17] protein folding and aggregation into amyloids. [18][19][20][21] Human lysozyme forms amyloid fibrils in vitro upon incubation at low pH and elevated temperature, 22,23 whereas HEWL forms amyloid fibrils when incubated in organic solvents 24 or in the presence of negatively charged lipid membranes.…”