title: Identification and biochemical characterization of the plasma membrane glucose transporter of Leishmania donovani.
abstract: The plasma membrane glucose transporter of Leishmania donovani, an obligate intracellular protozoan parasite of humans, was specifically labeled, identified, and biochemically characterized. Cytochalasin B, a known inhibitor of D-glucose transport in mammalian cells, but not cytochalasin E inhibited the transport of 2-deoxy-D-glucose in the extracellular promastigote form of this organism. Hydroxysuccinimidyl-4-azido-benzoate was used to photochemically cross-link [3H] cytochalasin B to the glucose transporter in isolated surface membranes and plasma membrane vesicles of L. donovani promastigotes. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the covalently labeled glucose transporter migrated as a 20-30-kDa protein band. This band was eluted from SDS-PAGE gels and subsequently analyzed by isoelectric focusing. The latter revealed two major peaks focusing at pH 6.8 and 6.6 [3H]Cytochalasin B-labeled membrane activity was detergent-solubilized, bound to concanavalin A-agarose beads, and specifically eluted with alpha-methyl mannoside. Analysis of the eluted material by SDS-PAGE revealed a D-glucose-inhibitable cytochalasin B peak with an apparent Mr approximately 20,000. The cumulative results indicate that the glucose transporter of L. donovani promastigotes is a glycoprotein which contains mannose as its major carbohydrate constituent.
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003571:Cytochalasin B / Q000494:pharmacology; D003847:Deoxyglucose / Q000378:metabolism; D007700:Kinetics; D007893:Leishmania donovani / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*
chemical_list: D009004:Monosaccharide Transport Proteins; D003571:Cytochalasin B; D003847:Deoxyglucose