Fig. 2: Prioritized Yck2 inhibitors selectively inhibit C. albicans Yck2.
a Purified C. albicans Yck2 kinase ___domain (0.115 µg/reaction) and human CK1α (0.05 µg/reaction) were treated in a two-fold dose response with each Yck2 inhibitor. Reactions were supplemented with dephosphorylated casein as substrate (2 µg) and ATP at the Km of the relevant kinase (CaYck2: 20 µM (closed circles), HsCK1α: 10 µM (open circles)). Following incubation, relative kinase activity was measured using the ADP-Glo™ (Promega) bioluminescent assay (100 ms integration). Data were normalized to compound-free controls, and IC50 values computed using GraphPad Prism 9, n = 3 (technical triplicates), Error bars represent the mean ± standard deviation (SD). The experiment was performed in at least a biological duplicate with similar results. b The susceptibility of a doxycycline-repressible tetO-YCK2/yck2Δ strain of C. albicans was evaluated in the presence and absence of doxycycline (10 µg/mL). Two-fold dose-response assays were performed in RPMI at 37 °C under 5% CO2 for 48 h. Growth was determined by optical density at 600 nm (OD600) with technical duplicate measurements averaged and normalized to compound-free controls (see color bar). The experiment was performed in biological duplicate with identical results. c The ability of each GW analog to potentiate caspofungin was tested in an echinocandin-resistant C. albicans strain (DPL15; FKS1T1922C/FKS1T1922C) using a two-fold dose-response assay in YPD at 30 ˚C for 48 h in the presence or absence of a sub-inhibitory concentration of caspofungin (3 µg/mL). Growth was determined by optical density at 600 nm (OD600) with technical duplicate measurements averaged and normalized to compound-free controls (see color bar). The experiment was performed in biological duplicate with identical results. Source data are provided as a Source Data file.
