The objectives of this project are: (1) A study of the kinetics and extent of vesicle-vesicle fusion in (i) pure and mixed phospholipid vesicles and (ii) vesicles containing glycolipids. This study will characterize the process of membrane fusion as a function of composition and size of vesicles, temperature, ion concentrations (Ca++, Mg2+, La3+, Na+), and effect of the fusogen polyethylene glycol. The long term objective is clarification of the mechanisms of membrane fusion. (2) Classification of factors affecting fusion of phospholipid vesicles with cells and analysis of the optimal conditions for capture and storage of molecules in vesicles and their subsequent transfer into target cells. (3) Analysis of the kinetics and equilibrium distribution of vesicle aggregation and their dependence on vesicle concentration and size, specifications, ligands and temperature. The study will focus on guiding experimental work by theoretical analysis of the mass action model for the kinetics and equilibrium of both aggregation and fusion of vesicles. Long-range intermolecular interactions will be calculated by further developing electrostatic expressions for free energies which take explicit account of cation binding to membranes. Short-range intermolecular interactions will be treated using phenomenological approaches and continuum field calculations, which consider the many-body effects of the environment. Collaborative experimental and theoretical studies will include 90 degree light scattering of aggregating and fusing vesicles and analysis of size changes; fluorescent intensity measurements of a Tb3+ complex formed in the interior of fusing vesicles; leakage measurements based on fluorescent intensity measurements of encapsulated carboxyfluorescein; leakage measurements of radio-labeled sucrose encapsulated in vesicles; X-ray diffraction to measure interbilayer distances in multilamellar vesicles in various concentrations of monovalent cations, and nuclear magnetic resonance studies (23Na and 31P) of binding of cations.
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