FIG 3.

Phosphorylation of EhCoactosin at S147. (A) Identification of EhCoactosin S147-containing phosphopeptide. The coactosin S147 phosphopeptide (EHI_168340A) KAGGADYSFNTTS(phospho)N was identified via SEQUEST with XCorr = 3.31, and precursor ΔM (experimental m/z and theoretical m/z) = −0.23 ppm. The MS/MS spectrum is annotated by Scaffold. The primary amines at the N terminus and on the lysine residue are dimethyl labeled. (B) Increased abundance of EhCoactosin S147-containing phosphopeptide in EhMSP-1-silenced Entamoeba histolytica. The MS1 spectrum of the light- (dimethyl-d0) and heavy-labeled (dimethyl-d6) EhCoactosin S147-containing phosphopeptides and the extracted ion chromatograms showing the light and heavy isotopologues eluting symmetrically at the same retention time are shown on the left and right, respectively. Heavy/light (H/L) was quantified by precursor ion elution profiles to be an average of 1.93 from multiple measurements from both biological replicates (one representative chromatograph [H/L = 1.98] is shown).