"Protein synthesis in rabbit reticulocytes: Characteristics of the prot" by Michael Grace, Robert O. Ralston et al.

Department of Chemistry

 

Document Type

Article

Date of this Version

1982

Comments

Published in Proc. NatL Acad. Sci. USA Vol. 79, pp. 6517-6521, November 1982. Used by Permission

Abstract

During heme deficiency in reticulocyte lysates, the heme-regulated translational inhibitor of protein synthesis (HRI) is activated and shuts off protein synthesis. In partial reactions, HRI phosphorylates the Mr 38,000 subunit (α subunit) of eukaryotic initiation factor 2 (eIF-2), which forms a ternary complex, Met-tRNAf•eIF-2•GTP. The eIF-2α(P) thus formed is not recognized by two eIF-2 ancillary factors, Co-eIF-2B (which promotes the dissociation of the ternary complex at high Mg2+) and Co-eIF-2C (which reverses the inhibition of ternary complex formation), and thus, is presumably inactive in peptide chain initiation. A protein factor, designated RF, which reverses inhibition of protein synthesis in heme-deficient reticulocyte lysates, has been purified -from reticulocyte cell supernatant. RF is a high molecular weight (Mr 450,000) protein complex composed of multiple polypeptides. An active RF preparation contains Co-eIF- 2B and Co-eIF-2C activities, and these two activities in RF preparation are not inhibited by HRI and ATP-i.e., eIF-2α(P) is recognized. During purification, RF remains associated with eIF-2 activity (eIF-2-RF) and can be freed of this eIF-2 activity by CMSephadex chromatography. Both eIF-2 RF and RF contain a Mr 38,000 polypeptide component that is indistinguishable from the Mr 38,000 subunit of eIF-2 by two-dimensional gel electrophoresis, It has been observed that a significant part of this Mr 38,000 polypeptide component in eIF-2 RF and almost the entire Mr 38,000 polypeptide component in RF remain unphosphorylated after prolonged incubation with HRI and ATP. A possible role of this free Mr 38,000 polypeptide in RF action is discussed.

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