A central issue in immunology concerns the understanding of the functional interaction between H2 coded antigens and neoantigens (i.e., viral tumor, or minorH). The problem has far- reaching implications in the mechanisms of immunity; the understanding of receptor specificity will shed light on T and B cell outogeny, function, and pathology. Cytotoxic T lymphocytes (CTLs) directed against the major surface glycoprotein (G protein) of vesicular stomatitis virus or the hemagglutinin (HA) of influenza virus recognize the viral protein in an H-2 restricted fashion. These viral antigens and H-2 proteins can be purified and incorporated into synthetic phospholipid membranes (liposomes) which are capable of stimulating a secondary CTL response. Not only is this response H-2 restricted, but the magnitude of the response is dependent on the H-2 haplotype. The goal of this project is to identify and characterize associations which occur between these viral antigens and H-2 proteins in liposomal membranes and to compare these associations to the biological reactivity of these liposomes. Saturation transfer electron paramagnetic reasonance spectroscopy and fluorescence energy transfer will be used to quantitate the interactions between viral and H-2 proteins in the liposomes. Further experiments designed to strengthen the correlation between membrane protein-protein association and CTL stimulation involve perturbation of the protein-protein interactions by several techniques. The degree of perturbation of these protein-protein associations will be measured by the spectroscopies mentioned above and the biological activity (CTL stimulation) of the perturbed samples will be determined. In addition, we will quantitate the extent of association between these viral proteins and H-2 molecules in virusinfected cells (these cells are sensitive to lysis by antiviral CTLs) by using fluorescentlabeled Fab fragments from monoclonal antibodies specific for the viral proteins and H-2 molecules. In this case fluorescence energy transfer will be measured using fluorescence flow cytometry. The longterm objective is to evaluate the molecular requirements for recognition of a determinant by the T cell receptor.
