The Crystal Structure of the N-terminal Region of the Alpha Subunit of Translation Initiation Factor 2 (eIF2α) from Saccharomyces cerevisiae Provides a View of the Loop Containing Serine 51, the Target of the eIF2α-specific Kinases

Dhaliwal, Simrit; Hoffman, David W.

doi:10.1016/j.jmb.2003.09.045

Cited by 53 publications

(57 citation statements)

References 38 publications

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“…1 A) (18). Structural analyses of free eIF2␣ indicate that, although the helix insert can adopt a precise ordered conformation, it is likely to be inherently flexible in nature [as reflected by high relative B-factors (18,19)]. This is consistent with the observation that the helix insert region of eIF2␣ was dynamically disordered in the PKR-eIF2␣ complex structure (12).…”

Section: Discussionsupporting

confidence: 65%

“…The helix insert region in eIF2␣ consists of 2 short 3/10 helices separated by a linker containing the phospho-accepting Ser-51 residue (Fig. 1 A) (18). Structural analyses of free eIF2␣ indicate that, although the helix insert can adopt a precise ordered conformation, it is likely to be inherently flexible in nature [as reflected by high relative B-factors (18,19)].…”

Section: Discussionmentioning

confidence: 99%

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Protein kinase PKR mutants resistant to the poxvirus pseudosubstrate K3L protein

Seo

Liu

Kawagishi-Kobayashi

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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As part of the mammalian cell innate immune response, the doublestranded RNA activated protein kinase PKR phosphorylates the translation initiation factor eIF2␣ to inhibit protein synthesis and thus block viral replication. Poxviruses including vaccinia and smallpox viruses express PKR inhibitors such as the vaccinia virus K3L protein that resembles the N-terminal substrate-targeting domain of eIF2␣. Whereas high-level expression of human PKR was toxic in yeast, this growth inhibition was suppressed by coexpression of the K3L protein.We used this yeast assay to screen for PKR mutants that are resistant to K3L inhibition, and we identified 12 mutations mapping to the C-terminal lobe of the PKR kinase domain. The PKR mutations specifically conferred resistance to the K3L protein both in yeast and in vitro. Consistently, the PKR-D486V mutation led to nearly a 15-fold decrease in K3L binding affinity yet did not impair eIF2␣ phosphorylation. Our results support the identification of the eIF2␣-binding site on an extensive face of the C-terminal lobe of the kinase domain, and they indicate that subtle changes to the PKR kinase domain can drastically impact pseudosubstrate inhibition while leaving substrate phosphorylation intact. We propose that these paradoxical effects of the PKR mutations on pseudosubstrate vs. substrate interactions reflect differences between the rigid K3L protein and the plastic nature of eIF2␣ around the Ser-51 phosphorylation site.eIF2 ͉ translational control

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Section: Discussionsupporting

confidence: 65%

Section: Discussionmentioning

confidence: 99%

Protein kinase PKR mutants resistant to the poxvirus pseudosubstrate K3L protein

Seo

Liu

Kawagishi-Kobayashi

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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“…Structures of yeast (Dhaliwal and Hoffman 2003;Hussain et al 2014;Llacer et al 2015), archaeal (Schmitt et al 2012), and mammalian eIF2a (Ito et al 2004) revealed that the protein consists of three domains: an N-terminal OB-fold domain and a central a-helical domain that are connected through a flexible linker to a C-terminal a/b domain that binds to eIF2g ( Figure 3A) (Schmitt et al 2012;Hussain et al 2014;Llacer et al 2015). A key mode of translational control in yeast and other eukaryotes involves phosphorylation of eIF2a.…”

Section: Ternary Complex Formationmentioning

confidence: 99%

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

2016

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In this review, we provide an overview of protein synthesis in the yeast Saccharomyces cerevisiae. The mechanism of protein synthesis is well conserved between yeast and other eukaryotes, and molecular genetic studies in budding yeast have provided critical insights into the fundamental process of translation as well as its regulation. The review focuses on the initiation and elongation phases of protein synthesis with descriptions of the roles of translation initiation and elongation factors that assist the ribosome in binding the messenger RNA (mRNA), selecting the start codon, and synthesizing the polypeptide. We also examine mechanisms of translational control highlighting the mRNA cap-binding proteins and the regulation of GCN4 and CPA1 mRNAs.

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“…The Ser51 phosphorylation site is located in domain 1 of eIF2α; however, mutational analyses of eIF2α revealed that substrate recognition by the eIF2α kinases, as well as binding of eIF2α to eIF2B, requires residues that are remote from the phosphorylation site (31,32). Moreover, biochemical and structural studies revealed that the two N-terminal domains of eIF2α assemble into a stable ∼200-residue protein and that further truncations lead to instability (30)(31)(32)(33)(34)(35). We anticipated that this N-terminal region of eIF2α might also be important for GADD34 recognition, although we could not rule out contributions from the eIF2α C-terminal domain.…”

Section: Viral Targeting Subunits Of Pp1 Promote Eif2α Dephosphorylatmentioning

confidence: 99%

An eIF2α-binding motif in protein phosphatase 1 subunit GADD34 and its viral orthologs is required to promote dephosphorylation of eIF2α

Rojas

Vasconcelos

Dever

2015

Proc. Natl. Acad. Sci. U.S.A.

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Transient protein synthesis inhibition, mediated by phosphorylation of the α subunit of eukaryotic translation initiation factor 2 (eIF2α), is an important protective mechanism cells use during stress conditions. Following relief of the stress, the growth arrest and DNA damage-inducible protein GADD34 associates with the broadly acting serine/threonine protein phosphatase 1 (PP1) to dephosphorylate eIF2α. Whereas the PP1-binding motif on GADD34 has been defined, it remains to be determined how GADD34 directs PP1 to specifically dephosphorylate eIF2α. In this report, we map a novel eIF2α-binding motif to the C terminus of GADD34 in a region distinct from where PP1 binds to GADD34. This motif is characterized by the consensus sequence Rx [Gnl], where capital letters are preferred and x is any residue. Point mutations altering the eIF2α-binding motif impair the ability of GADD34 to interact with eIF2α, promote eIF2α dephosphorylation, and suppress PKR toxicity in yeast. Interestingly, this eIF2α-docking motif is conserved among viral orthologs of GADD34, and is necessary for the proteins produced by African swine fever virus, Canarypox virus, and Herpes simplex virus to promote eIF2α dephosphorylation. Taken together, these data indicate that GADD34 and its viral orthologs direct specific dephosphorylation of eIF2α by interacting with both PP1 and eIF2α through independent binding motifs.

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The Crystal Structure of the N-terminal Region of the Alpha Subunit of Translation Initiation Factor 2 (eIF2α) from Saccharomyces cerevisiae Provides a View of the Loop Containing Serine 51, the Target of the eIF2α-specific Kinases

Cited by 53 publications

References 38 publications

Protein kinase PKR mutants resistant to the poxvirus pseudosubstrate K3L protein

Protein kinase PKR mutants resistant to the poxvirus pseudosubstrate K3L protein

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

An eIF2α-binding motif in protein phosphatase 1 subunit GADD34 and its viral orthologs is required to promote dephosphorylation of eIF2α

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