Figure 1.

Has1 is a component of both small and large pre-ribosomal particles. (A) An SDS PAGE analysis of the proteins co-purified with the wild type Has1-FTP. The TEV (10%) and FLAG eluates were resolved in 4–12% gradient SDS-PAGE and stained by Colloidal Coomassie Blue. The bait protein Has1 is marked with the asterisk. The marked proteins were identified by MALDI-TOF mass spectrometry. Pre-60S factors are depicted in blue and 90S/pre-40S factors in green. (B) The protein composition of pre-ribosomes purified via the wild type Has1-FTP. The iBAQ values normalized to bait are plotted (Has1 value was fixed to 10⁶). (C) Northern blotting analysis of RNA composition of pre-ribosomes affinity purified via plasmid-borne Has1-FTP (wild type or mutant). The endogenous Has1 wild type was not depleted. (D) Proteins with at least 2-fold change in their abundance in the pre-ribosomes purified via the catalytic mutants of Has1. The data from Has1-KA, Has1-DAAD and Has1-AAA experiments were pooled and the average SILAC ratios are shown. Only proteins present in pre-ribosomes from all three mutants were considered. The SILAC H/L ratios were normalized to bait. The error bars represent the standard deviation from the mean H/L ratio of the mutants across experiments.