FIG 5.

CK1 represents a bulk of the total kinase activity. (A) Kinase assays using 27 μM CK-S and 20 μg of total protein extract from promastigotes or axenic amastigotes were performed in the presence of either 1% DMSO or IC261, D4476, and CKI-7 at 10 μM in 1% DMSO. The activities in the reaction mixtures, spotted on P81 paper, were measured using a scintillation counter. For IC261 and CKI-7, there were no statistically significant differences between the inhibition obtained in promastigotes or in amastigotes (P = 0.510 and P = 0.111, respectively). There were no statistically significant differences between DMSO and IC261 (P = 0.573) and DMSO and CKI-7 (P = 0.286) in promastigotes or between DMSO and IC261 (P = 0.237) in amastigotes, but there was a significant difference between DMSO and CKI-7 (P < 0.001) in amastigotes. (B) Endogenous phosphorylation of 20 μg total protein extract from promastigotes (lanes P) and axenic amastigotes (lanes A) in the presence of either 1% DMSO or D4476, IC261, and CKI-7 at 10 μM in 1% DMSO. The reactions were analyzed by SDS-PAGE, and the gels were stained by colloidal Coomassie for loading assessment (top) and autoradiography (bottom). Quantification was estimated by the quantitative imaging software ImageJ.