Fig. 2: Identification of the P. falciparum target of OSM-S-106.

A P. falciparum cultures (Cam3.II-rev; trophozoite stage; 30–35 h p.i.) were exposed to OSM-S-106 for 6 h. Protein translation was assessed in the last 2 h of the incubation, via the incorporation of OPP. Aliquots of inhibitor-exposed cultures were washed and returned to cultures, and viability was estimated at the trophozoite stage of the next cycle. IC₅₀ (Translation) = 0.51 μM, IC₅₀ (Viability) = 4.7 μM. Error bars correspond to SEM of three independent experiments. B Trophozoite stage Cam3.II_rev parasites (30–35 h p.i.) were incubated with 0.05% DMSO (Mock), 50 nM borrelidin (BOR) or 2.5 μM OSM-S-106 or 2.5 μM OSM-S-137 for 3 h. Western blots of lysates were probed for phosphorylated eIF2α with PfBiP as a loading control. The blot is representative of three independent experiments and additional blots are presented in Supplementary Fig. 1C. C Schematic showing aaRS-catalyzed attack of a nucleoside sulfamate on an activated amino acid to form an amino acid adduct. D Structure of Asn-OSM-S-106. E P. falciparum-infected RBCs were treated with 10 μM OSM-S-106 for 3 h. Extracts were subjected to LCMS. The extracted ion chromatograms of the Asn-OSM-S-106 adduct generated by P. falciparum (upper panel) and the synthetic conjugate at m/z 421.0753 (lower panel). The inset shows MS analysis of the parasite-generated Asn-OSM-S-106 adduct. F Sensitivity to OSM-S-106 exposure (72 h) for a cloned wildtype line (Dd2) and a CRISPR-edited clone harboring PfAsnRS^R487S. Data represent five replicates and error bars correspond to SD. See Supplementary Table 6 for data values. Sensitivity to OSM-S-106 exposure (72 h) for aptamer-regulatable PfAsnRS (G) and PfNT4 (H) lines upon addition of aTc (closed circles) and with the target expression reduced (open circles), with data normalized to a no drug control. Data represent the mean of three replicates and error bars correspond to SD. See Supplementary Table 7 for data values.