Interaction of fMet‐tRNA<sup>fMet</sup> with the C‐terminal domain of translational initiation factor IF2 from <i>Bacillus stearothermophilus</i>

Krafft, Christoph; Diehl, Anne; Laettig, Stefan; Behlke, Joachim; Heinemann, Udo; Pon, Cynthia L.; Gualerzi, Claudio O.; Welfle, Heinz

doi:10.1016/s0014-5793(00)01377-6

Cited by 24 publications

(20 citation statements)

References 27 publications

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“…Domain IV of IF2, the bacterial ortholog of eIF5B, binds directly to the fMet portion of fMet-tRNA f Met Krafft et al 2000;Spurio et al 2000;Szkaradkiewicz et al 2000). Whereas eIF5B has not been shown to directly and stably bind Met-tRNA i Met , eIF5B has been found to substitute for eIF2 and promote Met-tRNA i Met binding during translation of certain viral mRNAs (Pestova et al 2008;Terenin et al 2008).…”

Section: Discussionmentioning

confidence: 99%

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Shin

Acker

Kim

et al. 2011

RNA

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Translation initiation factor eIF5B promotes GTP-dependent ribosomal subunit joining in the final step of the translation initiation pathway. The protein resembles a chalice with the α-helix H12 forming the stem connecting the GTP-binding domain cup to the domain IV base. Helix H12 has been proposed to function as a rigid lever arm governing domain IV movements in response to nucleotide binding and as a molecular ruler fixing the distance between domain IV and the G domain of the factor. To investigate its function, helix H12 was lengthened or shortened by one or two turns. In addition, six consecutive residues in the helix were substituted by Gly to alter the helical rigidity. Whereas the mutations had minimal impacts on the factor's binding to the ribosome and its GTP binding and hydrolysis activities, shortening the helix by six residues impaired the rate of subunit joining in vitro and both this mutation and the Gly substitution mutation lowered the yield of Met-tRNA(i)(Met) bound to 80S complexes formed in the presence of nonhydrolyzable GTP. Thus, these two mutations, which impair yeast cell growth and enhance ribosome leaky scanning in vivo, impair the rate of formation and stability of the 80S product of subunit joining. These data support the notion that helix H12 functions as a ruler connecting the GTPase center of the ribosome to the P site where Met-tRNA(i)(Met) is bound and that helix H12 rigidity is required to stabilize Met-tRNA(i)(Met) binding.

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Section: Discussionmentioning

confidence: 99%

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Shin

Acker

Kim

et al. 2011

RNA

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“…The Cterminal domain VI-2 of IF2 has been suggested to contain the entire binding site for fMet-tRNA f Met (215). The interaction of the domain with fMet-tRNA f Met has been studied using mutagenesis as well as Raman and NMR spectroscopy (66,96,130,225). Functionally essential residues of the B. stearothermophilus domain are C668, K699, R700, Y701, K702, E713, C714, and G715 (Fig.…”

Section: Bacillus Subtilis Is the Only Organism That Does Not Belong mentioning

confidence: 99%

Initiation of Protein Synthesis in Bacteria

Laursen

Sørensen

Mortensen

et al. 2005

Microbiol Mol Biol Rev

538

506

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Valuable information on translation initiation is available from biochemical data and recently solved structures. We present a detailed description of current knowledge about the structure, function, and interactions of the individual components involved in bacterial translation initiation. The first section describes the ribosomal features relevant to the initiation process. Subsequent sections describe the structure, function, and interactions of the mRNA, the initiator tRNA, and the initiation factors IF1, IF2, and IF3. Finally, we provide an overview of mechanisms of regulation of the translation initiation event. Translation occurs on ribonucleoprotein complexes called ribosomes. The ribosome is composed of a large subunit and a small subunit that hold the activities of peptidyltransfer and decode the triplet code of the mRNA, respectively. Translation initiation is promoted by IF1, IF2, and IF3, which mediate base pairing of the initiator tRNA anticodon to the mRNA initiation codon located in the ribosomal P-site. The mechanism of translation initiation differs for canonical and leaderless mRNAs, since the latter is dependent on the relative level of the initiation factors. Regulation of translation occurs primarily in the initiation phase. Secondary structures at the mRNA ribosomal binding site (RBS) inhibit translation initiation. The accessibility of the RBS is regulated by temperature and binding of small metabolites, proteins, or antisense RNAs. The future challenge is to obtain atomic-resolution structures of complete initiation complexes in order to understand the mechanism of translation initiation in molecular detail

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“…This module is structurally homologous to domain II of EF-G and EF-Tu (Nissen et al 1995;Brock et al 1998) and to the carboxy-terminal-most domain of IF2 itself, called IF2 C-2 . Following the GII domain, is C-1, a rather sturdy domain rich in helical structures (Misselwitz et al 1997;Krafft et al 2000;Spurio et al 2000). The last 110 amino acids of the protein (from Glu 632 to Ala 741) constitute the C-2 domain, which is responsible for the recognition and binding of fMet-tRNA Krafft et al 2000;Spurio et al 2000).…”

Section: Experimental Strategymentioning

confidence: 99%

“…Following the GII domain, is C-1, a rather sturdy domain rich in helical structures (Misselwitz et al 1997;Krafft et al 2000;Spurio et al 2000). The last 110 amino acids of the protein (from Glu 632 to Ala 741) constitute the C-2 domain, which is responsible for the recognition and binding of fMet-tRNA Krafft et al 2000;Spurio et al 2000).…”

Section: Experimental Strategymentioning

confidence: 99%

Ribosomal localization of translation initiation factor IF2

Marzi¹,

Knight²,

Brandi³

et al. 2003

RNA

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Bacterial translation initiation factor IF2 is a GTP-binding protein that catalyzes binding of initiator fMet-tRNA in the ribosomal P site. The topographical localization of IF2 on the ribosomal subunits, a prerequisite for understanding the mechanism of initiation complex formation, has remained elusive. Here, we present a model for the positioning of IF2 in the 70S initiation complex as determined by cleavage of rRNA by the chemical nucleases Cu(II):1,10-orthophenanthroline and Fe(II):EDTA tethered to cysteine residues introduced into IF2. Two specific amino acids in the GII domain of IF2 are in proximity to helices H3, H4, H17, and H18 of 16S rRNA. Furthermore, the junction of the C-1 and C-2 domains is in proximity to H89 and the thiostrepton region of 23S rRNA. The docking is further constrained by the requisite proximity of the C-2 domain with P-site-bound tRNA and by the conserved GI domain of the IF2 with the large subunit's factor-binding center. Comparison of our present findings with previous data further suggests that the IF2 orientation on the 30S subunit changes during the transition from the 30S to 70S initiation complex.

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Interaction of fMet‐tRNA^fMet with the C‐terminal domain of translational initiation factor IF2 from Bacillus stearothermophilus

Cited by 24 publications

References 27 publications

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Initiation of Protein Synthesis in Bacteria

Ribosomal localization of translation initiation factor IF2

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Interaction of fMet‐tRNAfMet with the C‐terminal domain of translational initiation factor IF2 from Bacillus stearothermophilus

Cited by 24 publications

References 27 publications

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Structural integrity of α-helix H12 in translation initiation factor eIF5B is critical for 80S complex stability

Initiation of Protein Synthesis in Bacteria

Ribosomal localization of translation initiation factor IF2

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Interaction of fMet‐tRNA^fMet with the C‐terminal domain of translational initiation factor IF2 from Bacillus stearothermophilus