This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Protozoan parasites of the genus Leishmania cause a number of diseases in humans. The parasite undergoes a complex life cycle, alternating between an insect vector (promastigotes) and their mammalian hosts (amastigotes). In a mouse model, vaccination with proteoglycolipid complex (P-8) of Leishmania pifanoi amastigotes provides partial to complete protection against infection by this organism. Therefore the P-8 complex represents a potential vaccine candidate for leishmaniasis and structural characterization of the glycolipid antigens of Leishmania pifanoi amastigotes is a necessary step in the vaccine development. Monoclonal antibodies to the P-8 complex recognize four related glycolipid components, but detailed structures of the glycolipids in the active fractions remain to be determined. Based on the preliminary results, the glycolipids are expected to belong to one of the classes of either glycoinositol-phospholipids or lipophosphoglycan precursors. Monosaccharide composition analysis confirms the presence of carbohydrate components in each fraction. MALDI FTMS offers a number of advantages for structural analysis of glycolipids such as direct ionization of samples retained on the surface of TLC plates and capability for multistage dissociation of MALDI-generated ions. Previous MALDI FTMS and MS/MS spectra obtained for the P8 samples as unseparated mixtures, as partly purified components and as spots separated on TLC, are consistent with the presence of glycolipids with carbohydrate moieties of varying lengths. So far, all the structures match types previously reported. ESI-MS has led to the identification of a few simple lipids, and MALDI-TOF MS has yielded previously-characterized m/z values, but not novel ones. Further purification combined with new improvements of our TLC MALDI FTMS capabilities, and a newly set up system for lipid LCMS should help in the acquisition of full structural information that will enable identification of minor components in the active fractions.
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