Year-end Sale % OFF 
Abstract
The protein kinases PKR, GCN2, and PERK phosphorylate translation initiation factor eIF2alpha to regulate general and genespecific protein synthesis under various cellular stress conditions. Recent x-ray crystallographic structures of PKR and GCN2 revealed distinct dimeric configurations of the kinase domains. Whereas PKR kinase domains dimerized in a back-to-back and parallel orientation, the GCN2 kinase domains displayed an antiparallel orientation. The dimerization interfaces on PKR and GCN2 were localized to overlapping surfaces on the N-terminal lobes of the kinase domains but utilized different intermolecular contacts. A key feature of the PKR dimerization interface is a salt bridge interaction between Arg(262) from one protomer and Asp(266) from the second protomer. Interestingly, these two residues are conserved in all eIF2alpha kinases, although in the GCN2 structure, the two residues are too remote to interact. To test the importance of this potential salt bridge interaction in PKR, GCN2, and PERK, the residues constituting the salt bridge were mutated either independently or together to residues with the opposite charge. Single mutations of the Asp (or Glu) and Arg residues blocked kinase function both in yeast cells and in vitro. However, for all three kinases, the double mutation designed to restore the salt bridge interaction with opposite polarity resulted in a functional kinase. Thus, the salt bridge interaction and dimer interface observed in the PKR structure is critical for the activity of all three eIF2alpha kinases. These results are consistent with the notion that the PKR structure represents the active state of the eIF2alpha kinase domain, whereas the GCN2 structure may represent an inactive state of the kinase.
Highlights
Protein synthesis in mammalian cells is regulated by four protein kinases (PKR, PERK, GCN2, and HRI) that phosphorylate the ␣ subunit of the translation initiation factor eIF2 in response to various environmental stress conditions
The isolated, kinase domain (KD), HisRS, and C-terminal domain of GCN2 were found to dimerize in vivo, and these dimer contacts are thought to maintain the protein in a dimeric state [13]
Reciprocal Exchange of Residues Arg599 and Glu603 Restores PERK Function in Vivo and in Vitro—Like PKR, high level expression of PERK in yeast lacking GCN2 results in phosphorylation of eIF2␣ and inhibition of cell growth [11]. Consistent with these previous results, we found that high level expression of FLAG- and His6-tagged C. elegans PERK from a galactose-inducible promoter impaired the growth of the gcn2⌬ strain H2557 (Fig. 2D, left, row 2) and resulted in significant phosphorylation of eIF2␣ on Ser51 (Fig. 2E, lane 2)
Summary
Protein synthesis in mammalian cells is regulated by four protein kinases (PKR, PERK, GCN2, and HRI) that phosphorylate the ␣ subunit of the translation initiation factor eIF2 in response to various environmental stress conditions (reviewed in Refs. 1 and 2–5). The salt bridge interaction and dimer interface observed in the PKR structure is conserved among PKR, GCN2, and PERK and is critical for eIF2␣ kinase activation.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
