Figure 4. Thermal proteome profiling (TPP) suggests PfHsp90-dependent processes.

(A) The substitution of a hydroxyl group in BX-2819 to a methoxyl group yields the inactive derivative HS292. (B) BX-2819 (10 μM) displaces FITC-GA, whereas HS292 (100 μM) does not compete for binding to PfHsp90. Data shown as means ± SEM, n = 5; not significant (ns)>0.05, ****<0.0001; one-way ANOVA, Dunnett’s multiple comparison. (C) Plasmodium proteins that did not exhibit a stability change (−2.0 °C < ΔTm < +2.0 °C; dotted line) upon treatment with the inactive HS292 but did exhibit a stability change (ΔTm < −2.0 °C or ΔTm > +2.0 °C; dotted line) upon treatment with the active BX-2819 inhibitor compared to DMSO were selected as TPP hits (orange fill indicates selection criteria for each subset). (D) Network of interactions between TPP hits. The PfHsp90 node (blue) represents both Q8IC05 (cytosolic) and Q8III6 (mitochondrial) parasite paralogs, whereas remaining nodes represent other TPP hit proteins with UniProt Identifiers. Edges connecting nodes represent predicted interactions between the proteins, with the line width corresponding to confidence. Only proteins within two edges of the PfHsp90 node are displayed. The significant local network cluster representing the proteasome (CL: 4475; false discovery rate: 0.0067) is highlighted in orange. (E) Gene ontology (GO) enrichment detailing biological processes of TPP hits. GO terms are scaled with the color representing the significance of the enriched annotations and the size representing the number of annotations for the ascribed process in the database.