CONICET | Buscador de Institutos y Recursos Humanos

In the field of signal transduction the importance of protein complex formation associated with limited regions of cell membranes is increasingly being recognized. These complexes form by means of protein-protein and protein-lipid interactions. The dynamic assembly and disassembly of these complexes frequently depend on postranslational modifications such as phosphorylation. Another modification proposed for membrane association is the S-palmitoylation, the covalent attachment of long chain fatty acids, principally palmitate, to cysteine residues. To evaluate the role of the S-palmitoylation in neuronal signal transduction, we have developed two approaches: (i) isolation of detergent resistant membrane (DRM) domains from synaptosomes of adult rat brain and (ii) protein expression in E. Coli with in vitro palmitoylation to analyze their interaction with artificial membranes. These two complementary approaches allow us to evaluate physiological factors involved in palmitoyl-protein translocation to complexes associated with neuronal synaptic membranes. In parallel, we can manipulate the conditions required for translocationin the experimentally defined liposome system. Thus we expressed and purified the protein GAP-43, and analyzed its association with artificial membranes and determined the contribution of S-palmitoylation to translocation to membrane domains. Comparing these two systems we can generate complementary information about the physiological regulation of the palmitoylation and the compartmentalizing of individual proteins.