The Re and Rd chemotypes of lipopolysaccharides (LPS) obtained from Escherichia coli and Salmonella minnesota will be derivatized to the methyl esters and purified to structural homogeneity by utilizing silica gel chromatography and the recently developed 50-nm pore size polyvinylbenzene gel permeation and divinylbenzene gel reverse-phase columns for high performance liquid chromatograhy (HPLC). These highly purified and structurally homogeneous methyl LPS will then be analyzed by chemical means, fast atom bombardment mass spectrometry, california plasma deserption mass spectrometry, 1-H-nuclear magnetic resonance (NMR) spectroscopy, and 31-p-NMR spectroscopy. Using these results, we shall attempt to establish the complete structures of the native LPS and determine the nature of their apparent microheterogeneit. The purified methyl LPS will be subjected to selective biological assays in order to study the LPS structure-to-biological-activity relationship. We shall examine how the biological activities of the lipid A moiety of purified methyl LPS containing 2-keto-3-deoxyoctonate might be modulated by the presence of one or more of the following polar groups: pyrophosphate, ethanolamine, aminoarabinose, and L-glycero-D-mannoheptose. These assays include tests for toxicity and antitumor activities. Lipid A from the following sources will be methylated and purified to homogeneity by HPLC: nontoxic diphosphoryl lipid A from Rhodopseudomonas sphaeroides, phosphatefree lipid A from Rhodomicrobium vannielii, and lipid A from Brucella abortus and Neisseria gonorrhoeae. Complete structures of these lipid A's will be determined and the underivatized samples will be tested for certain biological activities. The long-term objective of this research is to progress to the more complex Rc, Rb, and Ra LPS chemotypes in our structural studies and ultimately to be able to examine the structures of the highly purified smooth strain LPS at the intact level. Knowledge gained from this study might be applied to cancer immunotherapy, the development of an effective adjuvant, protection against X-irradiation, and protection against bacterial and viral infections.
Showing the most recent 10 out of 31 publications
