FIGURE 4:

PERK Y561F mutation enhances PERK catalytic activity. (A) Phosphorylation of recombinant His-eIF2α by GST-cPERK WT and Y561F mutant in in vitro kinase assay. Bar chart shows quantification by densitometry of His-eIF2α phosphorylation normalized for the amounts of eIF2α and GST-cPERK present in the reactions. Data are mean ± SEM (n = 3; *p < 0.01). (B) PERK^−/− MEFs transiently transfected with empty vector (Mock), full-length PERK WT, Y561F, or K618A cDNAs were treated 48 h later with 1 mM DTT for 0, 20, or 60 min. Cell lysates normalized for protein content (50 μg of protein) were subjected to immunoblotting with indicated antibodies. Shown are three independent experiments, with peIF2αS⁵¹/total eIF2α ratio, as determined by densitometry, reported under the blots.