Crystal structure of eukaryotic translation initiation factor 2B

Abstract

Eukaryotic cells restrict protein synthesis under various stress conditions, by inhibiting the eukaryotic translation initiation factor 2B (eIF2B). eIF2B is the guanine nucleotide exchange factor for eIF2, a heterotrimeric G protein consisting of α-, β- and γ-subunits. eIF2B exchanges GDP for GTP on the γ-subunit of eIF2 (eIF2γ), and is inhibited by stress-induced phosphorylation of eIF2α. eIF2B is a heterodecameric complex of two copies each of the α-, β-, γ-, δ- and ε-subunits; its α-, β- and δ-subunits constitute the regulatory subcomplex, while the γ- and ε-subunits form the catalytic subcomplex. The three-dimensional structure of the entire eIF2B complex has not been determined. Here we present the crystal structure of Schizosaccharomyces pombe eIF2B with an unprecedented subunit arrangement, in which the α2β2δ2 hexameric regulatory subcomplex binds two γε dimeric catalytic subcomplexes on its opposite sides. A structure-based in vitro analysis by a surface-scanning site-directed photo-cross-linking method identified the eIF2α-binding and eIF2γ-binding interfaces, located far apart on the regulatory and catalytic subcomplexes, respectively. The eIF2γ-binding interface is located close to the conserved 'NF motif', which is important for nucleotide exchange. A structural model was constructed for the complex of eIF2B with phosphorylated eIF2α, which binds to eIF2B more strongly than the unphosphorylated form. These results indicate that the eIF2α phosphorylation generates the 'nonproductive' eIF2-eIF2B complex, which prevents nucleotide exchange on eIF2γ, and thus provide a structural framework for the eIF2B-mediated mechanism of stress-induced translational control.

Structure of the nucleotide exchange factor eIF2B reveals mechanism of memory-enhancing molecule

“…Crosslinking studies suggest that eIF2 binds across the decameric interface, engaging the eIF2B α subunit, and β and δ subunits from opposing tetramers (8). We surmise that decamer assembly creates a composite surface for eIF2 binding that allows the flexibly attached HEAT domain to reach and engage its target.…”

“…eIF2B is composed of five subunits (α,β,γ,δ,ε) that assemble into a decamer composed of two copies of each subunit (4–8). The eIF2Bε subunit contains the enzyme’s catalytic center and associates closely with eIF2Bγ (9).…”

Fail-safe control of translation initiation by dissociation of eIF2α phosphorylated ternary complexes

“…This causes a general reduction in protein synthesis initiation rates while at the same time activating translation of ISR-responsive mRNAs (Young and Wek, 2016). Genetic and biochemical evidence shows that phosphorylated eIF2α binds to a regulatory site on eIF2B formed by the eIF2Bαβδ subcomplex (Pavitt et al, 1997; Krishnamoorthy et al, 2001; Kashiwagi et al, 2016b). In contrast GEF function is provided by eIF2Bε, which interacts with eIF2β and the GDP/GTP-binding eIF2γ subunit (Gomez and Pavitt, 2000; Asano et al, 1999; Alone and Dever, 2006).…”

Molecular Determinants of the Regulation of Development and Metabolism by Neuronal eIF2α Phosphorylation in Caenorhabditis elegans

“…The eif-2Ba mutations that confer the dominant-suppressor phenotype similar to the loss-of-function allele likely affect residues that are critical for eIF2Ba functional and structural integrity (Table S2 in File S1). Additionally, because eIF2Ba forms a homodimer as part of the eIF2B holoenzyme, the altered eIF2Ba may exert a dominant-negative effect that prevents proper dimerization or holoenzyme formation (Kashiwagi et al 2016).…”