Abstract
Malaria is a health burden disease caused by Plasmodium species, P. falciparum, P. vivax, P. P. ovale, malariae, and P. knowlesi. The most threatening one is P. falciparum. However, P. falciparum has become resistant to available antimalarial drugs. When the parasite attacks the red blood cell, proteins known as heat shock proteins (Hsp’s) are expressed and upregulated. The structure and function of these proteins make them potential targets for antimalarial drugs. Plasmodium falciparum Heat shock protein 90 (PfHsp90) is an upregulated protein during the erythrocyte stage of infection and works together with co-chaperones such as Plasmodium falciparum Heat shock protein 70 (PfHsp70) and Plasmodium falciparum Hsp70-Hsp90 organizing protein (PfHop) to facilitate client protein folding. Iso-mukaadial acetate (IMA) and Ursolic acid acetate (UAA) are active plant-derived compounds that have biological activity against Plasmodium species. This study aimed to evaluate the inhibitory activity of Iso-mukaadial acetate and Ursolic acid acetate against PfHsp90. PfHsp90 was successfully expressed in E. coli BL21 (DE3) and purified using nickel affinity chromatography. Ultraviolet-visible spectroscopy (UV-vis) and Fourier transform infrared spectroscopy (FTIR) were used to confirm the PfHsp90 secondary structural changes. UV-vis established that the PfHsp90 aromatic amino acids are buried within protein itself and IMA and UAA interact with aromatic amino acids rings to expose them. FTIR showed the interaction to PfHsp90 structurally compromises secondary structure elements where β-sheets and β-turns decreased, and α-helix and β-antiparallel increased so thus suggesting destabilization of PfHsp90. Chaperone activity of PfHsp90 and the inhibitory activity of study compounds were analyzed using malate dehydrogenase (MDH), luciferase, and Citrate synthase (CS) aggregation assays, which established that PfHsp90 suppresses aggregation while study compounds inhibit the PfHsp90 to suppress aggregation. Through basal ATPase activity assay, PfHsp90 was established that it hydrolyzes ATP at Km=138.8 μM. However, IMA and UAA binds to the N-terminal ATPase domain and inhibit PfHsp90 ATPase activity with Km=50.65 μM and 291.2 μM, respectively. In conclusion, IMA and UAA bind to PfHsp90, inhibit PfHsp90 chaperone activity and ATPase. This study expands the current knowledge in the biological activity of inhibitors against the molecular chaperone network...
M.Sc. (Biochemistry)