FIGURE 1.

Gcn2 co-elutes with endogenously expressed His₆-eEF1A. tef1Δ;tef2Δ double deletion strains expressing eEF1A from a plasmid from its own promotor, either untagged or His₆-tagged (TKY864 and TKY865, respectively), were grown to exponential phase and harvested. Whole cell extracts were incubated with iMAC resin, and the resins were then subsequently washed with increasing concentrations of imidazole as indicated. Equal amounts of each washing step were subjected to SDS-PAGE, and the proteins were transferred to a PVDF membrane. The membrane was probed with Ponceau S to visualize His₆-eEF1A and then was subjected to immunoblotting using antibodies against Gcn1, Gcn2, and the small ribosomal protein RPS22.