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TRANSLATIONAL LEVEL CONTROL OF PROTEIN SYNTHESIS IN EUKARYOTIC CELLS
Abstract
During heme-deficiency in reticulocyte lysate, a translational inhibitor (HRI, heme-regulated inhibitor) that blocks polypeptide chain initiation is activated. HRI is a protein kinase that specifically phosphorylates the 38,000-dalton subunit of the Met-tRNA(,f) binding factor, eIF-2. Phosphorylation of eIF-2 by HRI prevents its interaction with at least two additional factors resulting in a net reduction of ternary complex (Met-tRNA(,f)(.)eIF-2(.)GTP) formation and AUG-dependent transfer of Met-tRNA(,f) to 40S ribosomal subunits. A factor (RF) that reverses protein synthesis inhibition in heme-deficient lysates has been purified from reticulocyte ribosomal salt wash and post-ribosomal supernatant. The purified RF restores protein synthesis activity of the heme-deficient lysate to the level observed in the presence of hemin. No direct correlation exists between amount of eIF-2 activity and ability to reverse protein synthesis inhibition in heme-deficient lysates. Homogeneous preparations of eIF-2 are completely inactive in reversal of protein synthesis inhibition in heme-deficient lysate. RF also reverses the inhibition of ternary complex formation by HRI in a fractionated system. Both the ternary complex inhibition reversal and protein synthesis inhibition reversal activities co-sediment at 12.5S upon glycerol density gradient centrifugation and both activities are heat and NEM sensitive. Purified RF does not dephosphorylate phosphorylated eIF-2 (HRI-catalyzed) nor prevent the phosphorylation of eIF-2 by HRI in a fractionated system. RF stimulates ternary complex formation by both phosphorylated and non-phosphorylated eIF-2. These observations suggest that the sensitivity of protein synthesis to eIF-2 kinases such as HRI may be modulated by RF.
Subject Area
Biochemistry
Recommended Citation
RALSTON, ROBERT ORVILLE, "TRANSLATIONAL LEVEL CONTROL OF PROTEIN SYNTHESIS IN EUKARYOTIC CELLS" (1980). ETD collection for University of Nebraska-Lincoln. AAI8021352.
https://digitalcommons.unl.edu/dissertations/AAI8021352