Inhibition of glycosyl-phosphatidylinositol biosynthesis in Plasmodium falciparum by C-2 substituted mannose analogues.

Mannose analogues (2-deoxy-D-glucose, 2-deoxy-2-fluoro-D-glucose and 2-amino-2-deoxy-D-mannose) have been used to study glycosylphosphatidylinositol (GPtdIns) biosynthesis and GPtdIns protein anchoring in protozoal and mammalian systems. The effects of these analogues on GPtdIns biosynthesis and GPtdIns-protein anchoring of the human malaria parasite Plasmodium falciparum were evaluated in this study. At lower concentrations of 2-deoxy-D-glucose and 2-deoxy-2-fluoro-D glucose (0.2 and 0.1 mm, respectively), GPtdIns biosynthesis is inhibited without significant effects on total protein biosynthesis. At higher concentrations of 2-deoxy-D-glucose and 2-deoxy-2-fluoro-D-glucose (1.5 and 0.8 mm, respectively), the incorporation of [3H]glucosamine into glycolipids was inhibited by 90%, and the attachment of GPtdIns anchor to merozoite surface protein-1 (MSP-1) was prevented. However, at these concentrations, both sugar analogues inhibit MSP-1 synthesis and total protein biosynthesis. In contrast to 2-deoxy-2-fluoro-D-glucose and 2-amino-2-deoxy-D-mannose (mannosamine), the formation of new glycolipids was observed only in the presence of tritiated or nonradiolabelled 2-deoxy-D-glucose. Mannosamine inhibits GPtdIns biosynthesis at a concentration of 5 mm, but neither an accumulation of aberrant intermediates nor significant inhibition of total protein biosynthesis was observed in the presence of this analogue. Furthermore, the [3H]mannosamine-labelled glycolipid spectrum resembled the one described for [3H]glucosamine labelling. Total hydrolysis of mannosamine labelled glycolipids showed that half of the tritiated mannosamine incorporated into glycolipids was converted to glucosamine. This high rate of conversion led us to suggest that no actual inhibition from GPtdIns biosynthesis is achieved with the treatment with mannosamine, which is different to what has been observed for mammalian cells and other parasitic protozoa.