Lipooligosaccharides (LOSs) are important antigenic and pathogenic components of the outer membrane of Neisseria gonorrhoea. The long term goal of this project is to determine the stereochemistry of epitopes of gonococcal LOSs to understand the antigenicity and immunogenicity of the gonococcus. The gonococcal LOSs consist of heterogeneous components. Their epitope expression is heterogeneous, and such antigenic heterogeneity is due to molecular heterogeneity of each LOS preparation. Recent structural and immmunochemical characterization of LOS epitopes indicates that epitopes, defined by monoclonal antibodies, are within the carbohydrate moieties of LOS components. However, the oligosaccharide moieties must be in certain conformations for them to be expressed. Conformations of the OS moieties are defined by the overall conformations of the whole LOS components. Such organized LOS structures are significantly influenced by the lipoidal moieties that act as anchors for the carbohydrate moieties. We will further study structures and stereochemistry of LOS epitopes for their expression. We will use three monoclonal antibodies (two are IgM and one is IgG) to define epitopes in LOS molecules. These antibodies each identify separate LOS component and two of these antibodies bind to gonococcal strains in vivo. Therefore, characterization of their epitopes is very important. OS components from LOS samples will be analyzed by chemical methods and then further analyzed by several exo and endo glycosidases. Based on the above analysis, LOS components will be fragmented by sequential enzymatic treatments. The fragmented LOS components will be analyzed for their antigenic expression. Along with this analysis, we will characterize mono- and oligosaccharides, released by enzymatic treatments of OS and LOS components. OS components will also be analyzed by NMR before and after enzymatic treatments, to determine partial structures of intact OS components. Finally, based on the above studies, we will synthesize """"""""epitopes (OS- lipoidal moiety complexes)"""""""". Conformational analysis of antigenic epitopes will be carried out by NMR.
