Fig 8. WU-1 directly inhibits both the binding of the glucose analogue (ATB-BMPA) to PfHT and PfHT transport activity.

A) Biotinylated ATB-BMPA, a bis-mannose containing photolabel, binds to the glucose binding site of PfHT. Liposomes containing 0.65 μg PfHT were preincubated with ± 10 μM WU-1 prior to labeling with biotinylated ATB-BMPA. PfHT bound with biotinylated ATB-BMPA was analyzed by immunoblot analysis using a fluorescently labeled streptavidin (Strep). The strep signal was normalized to the amount of PfHT protein. Data are expressed as the means ± SEM., n = 3; (*), p<0.05 vs. vehicle control as determined by the paired Student’s t test. B) WU-1 inhibits the specific uptake ([³H]-D-glucose minus ([³H]-L-glucose) into PfHT-containing liposomes. Different concentrations of WU-1 were added to the liposomes 20 min prior to the initiation of the transport reaction. Uptake (quenched after 50 sec) was normalized to the amount of PfHT in the liposomes. Data were fit by nonlinear regression analysis using GraphPad Prism 6.0 software to calculate the IC₅₀ for WU-1. Data are expressed as mean ± SEM of three independent experiments.