The goal is to determine the stereochemistry of epitopes to Neisseria gonorrhoeae lipooligosaccharides (LOS) for the understanding of the immunobiology of the gonococcus, and this information will be important for the development of vaccines for gonococcal disease. The gonococcal LOS consists of oligosaccharide (OS) chains and a lipoidal moiety, called Lipid A. Antigenic expression within OS of the LOS is ruled by secondary or higher structures of the LOS, i.e., the epitope expression is the reflection of the fine structure of OS epitopes and they are expressed under such conditions that their stereochemical environment is fulfilled. We will focus on the stereochemical requirement of LOS epitopes for their expression. We will use two monoclonal antibodies (one IgG and one IgM) to define epitopes within two LOS for this project. LOS will be modified and also fragmented in various ways, and modified and fragmented LOS will be analyzed by chemical and immunochemical methods to determine the minimum size and structural requirement for the expression of the epitopes recognized by the two antibodies. OS components derived from antigenic fragmented LOS will be purified by chromatography (open column chromatography and HPLC) and their structures will be fully characterized by GC, GC/MS, FAB/MS and NMR spectroscopy. Those OS components will be coupled to hydrophobic substances, and these complexes will be immunochemically analyzed to understand the role and function of lipid A moiety for the expression of epitopes. Finally, conformational aspects of epitopes will be studied by NMR and CD using fragmented LOS, OS, and OS complexes. This analysis will also be supported by comparative studies of synthesized, small oligosaccharides.
