The proposed project is directed to develop a quantitative characterization, at the molecular level, of the structural organization and functional properties of glycolipid receptors, namely gangliosides. These studies will be performed on intact synaptosomal membranes from brain tissue as well as phospholipid vesicles containing well defined concentrations of purified gangliosides. We will examine the lateral distribution of gangliosides in synaptosomal membranes and liposomes as well as their lateral mobility and rotational mobility using steady state and time resolved fluorescence and phosphorescence spectroscopy. Conformational transitions of proteins and lipid phase transitions will be examined in detail using high sensitivity differential scanning calorimetry and isothermal microcalorimetry. The resulting characterization will be correlated with ganglioside functional parameters. The enzymatic activity of soluble neuraminidases as well as synaptosomal membrane bound neuraminidase will be evaluated as a function of the composition, structure and physical state of the membrane. The sequence of events characterizing the binding, insertion and lateral redistribution of molecules following the association of Cholera Toxin to membrane systems containing ganglioside GM1 will be evaluated in detail. These studies will provide fundamental information about the role of gangliosides as major constituents of central nervous tissue, as membrane surface receptors and as membrane bound enzyme substrates, and the factors affecting their physiological function. In this way the development of this project will contribute to our understanding of the molecular basis of cellular membrane alterations found in ganglioside storage diseases.
