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Emmanuelle Schmitt - One of the best experts on this subject based on the ideXlab platform.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation
Communications Biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 Å resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation.
Communications biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 A resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life. Coureux et al. describe a cryo-EM structure of a 30S initiation complex of Pyrococcus abyssi at 3.2A resolution. The structure uncovers a novel archaeal ribosomal protein aS21, N-terminal extension of eL41 and brings insights into base modifications of the rRNA.
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Start Codon Recognition in Eukaryotic and Archaeal Translation Initiation: A Common Structural Core
International Journal of Molecular Sciences, 2019Co-Authors: Emmanuelle Schmitt, Etienne Dubiez, Auriane Monestier, Pierre-damien Coureux, Yves MechulamAbstract:Understanding molecular mechanisms of ribosomal translation sheds light on the emergence and evolution of protein synthesis in the three domains of life. Universally, ribosomal translation is described in three steps: initiation, elongation and termination. During initiation, a macromolecular complex assembled around the small ribosomal subunit selects the start codon on the mRNA and defines the open reading frame. In this review, we focus on the comparison of start codon selection mechanisms in eukaryotes and archaea. Eukaryotic translation initiation is a very complicated process, involving many initiation factors. The most widespread mechanism for the discovery of the start codon is the scanning of the mRNA by a pre-initiation complex until the first AUG codon in a correct context is found. In archaea, long-range scanning does not occur because of the presence of Shine-Dalgarno (SD) sequences or of short 5' untranslated regions. However, archaeal and eukaryotic translation initiations have three initiation factors in common: e/AIF1, e/AIF1A and e/aIF2 are directly involved in the selection of the start codon. Therefore, the idea that these archaeal and eukaryotic factors fulfill similar functions within a common structural ribosomal core complex has emerged. A divergence between eukaryotic and archaeal factors allowed for the adaptation to the long-range scanning process versus the SD mediated prepositioning of the ribosome.
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Role of AIF1 in Pyrococcus abyssi translation initiation
Nucleic Acids Research, 2018Co-Authors: Auriane Monestier, Christine Lazennec-Schurdevin, Yves Mechulam, Pierre-damien Coureux, Emmanuelle SchmittAbstract:In archaeal translation initiation, a preinitiation complex (PIC) made up of AIF1, AIF1A, the ternary complex (TC, e/aIF2-GTP-Met-tRNA i Met) and mRNA bound to the small ribosomal subunit is responsible for start codon selection. Many archaeal mRNAs contain a Shine-Dalgarno (SD) sequence allowing the PIC to be prepositioned in the vicinity of the start codon. Nevertheless, cryo-EM studies have suggested local scanning to definitely establish base pairing of the start codon with the tRNA anticodon. Here, using flu-orescence anisotropy, we show that AIF1 and mRNA have synergistic binding to the Pyrococcus abyssi 30S. Stability of 30S:mRNA:AIF1 strongly depends on the SD sequence. Further, toeprinting experiments show that AIF1-containing PICs display a dynamic conformation with the tRNA not firmly accommodated in the P site. AIF1-induced destabilization of the PIC is favorable for proofreading erroneous initiation complexes. After AIF1 departure, the stability of the PIC increases reflecting initiator tRNA fully base-paired to the start codon. Altogether, our data support the idea that some of the main events governing start codon selection in eukaryotes and archaea occur within a common structural and functional core. However, idiosyncratic features in loop 1 sequence involved in 30S:mRNA binding suggest adjustments of e/AIF1 functioning in the two domains.
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Identification of a second GTP-bound magnesium ion in archaeal initiation factor 2
Nucleic Acids Research, 2016Co-Authors: Etienne Dubiez, Christine Lazennec-Schurdevin, Alexey Aleksandrov, Yves Mechulam, Emmanuelle SchmittAbstract:Eukaryotic and archaeal translation initiation processes involve a heterotrimeric GTPase e/aIF2 crucial for accuracy of start codon selection. In eu-karyotes, the GTPase activity of eIF2 is assisted by a GTPase-activating protein (GAP), eIF5. In ar-chaea, orthologs of eIF5 are not found and aIF2 GT-Pase activity is thought to be non-assisted. However , no in vitro GTPase activity of the archaeal factor has been reported to date. Here, we show that aIF2 significantly hydrolyses GTP in vitro. Within aIF2␥, H97, corresponding to the catalytic histidine found in other translational GTPases, and D19, from the GKT loop, both participate in this activity. Several high-resolution crystal structures were determined to get insight into GTP hydrolysis by aIF2␥. In particular, a crystal structure of the H97A mutant was obtained in the presence of non-hydrolyzed GTP. This structure reveals the presence of a second magnesium ion bound to GTP and D19. Quantum chemical/molecular mechanical simulations support the idea that the second magnesium ion may assist GTP hydrolysis by helping to neutralize the developing negative charge in the transition state. These results are discussed in light of the absence of an identified GAP in archaea to assist GTP hydrolysis on aIF2.
Yves Mechulam - One of the best experts on this subject based on the ideXlab platform.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation
Communications Biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 Å resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation.
Communications biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 A resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life. Coureux et al. describe a cryo-EM structure of a 30S initiation complex of Pyrococcus abyssi at 3.2A resolution. The structure uncovers a novel archaeal ribosomal protein aS21, N-terminal extension of eL41 and brings insights into base modifications of the rRNA.
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Start Codon Recognition in Eukaryotic and Archaeal Translation Initiation: A Common Structural Core
International Journal of Molecular Sciences, 2019Co-Authors: Emmanuelle Schmitt, Etienne Dubiez, Auriane Monestier, Pierre-damien Coureux, Yves MechulamAbstract:Understanding molecular mechanisms of ribosomal translation sheds light on the emergence and evolution of protein synthesis in the three domains of life. Universally, ribosomal translation is described in three steps: initiation, elongation and termination. During initiation, a macromolecular complex assembled around the small ribosomal subunit selects the start codon on the mRNA and defines the open reading frame. In this review, we focus on the comparison of start codon selection mechanisms in eukaryotes and archaea. Eukaryotic translation initiation is a very complicated process, involving many initiation factors. The most widespread mechanism for the discovery of the start codon is the scanning of the mRNA by a pre-initiation complex until the first AUG codon in a correct context is found. In archaea, long-range scanning does not occur because of the presence of Shine-Dalgarno (SD) sequences or of short 5' untranslated regions. However, archaeal and eukaryotic translation initiations have three initiation factors in common: e/AIF1, e/AIF1A and e/aIF2 are directly involved in the selection of the start codon. Therefore, the idea that these archaeal and eukaryotic factors fulfill similar functions within a common structural ribosomal core complex has emerged. A divergence between eukaryotic and archaeal factors allowed for the adaptation to the long-range scanning process versus the SD mediated prepositioning of the ribosome.
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Role of AIF1 in Pyrococcus abyssi translation initiation
Nucleic Acids Research, 2018Co-Authors: Auriane Monestier, Christine Lazennec-Schurdevin, Yves Mechulam, Pierre-damien Coureux, Emmanuelle SchmittAbstract:In archaeal translation initiation, a preinitiation complex (PIC) made up of AIF1, AIF1A, the ternary complex (TC, e/aIF2-GTP-Met-tRNA i Met) and mRNA bound to the small ribosomal subunit is responsible for start codon selection. Many archaeal mRNAs contain a Shine-Dalgarno (SD) sequence allowing the PIC to be prepositioned in the vicinity of the start codon. Nevertheless, cryo-EM studies have suggested local scanning to definitely establish base pairing of the start codon with the tRNA anticodon. Here, using flu-orescence anisotropy, we show that AIF1 and mRNA have synergistic binding to the Pyrococcus abyssi 30S. Stability of 30S:mRNA:AIF1 strongly depends on the SD sequence. Further, toeprinting experiments show that AIF1-containing PICs display a dynamic conformation with the tRNA not firmly accommodated in the P site. AIF1-induced destabilization of the PIC is favorable for proofreading erroneous initiation complexes. After AIF1 departure, the stability of the PIC increases reflecting initiator tRNA fully base-paired to the start codon. Altogether, our data support the idea that some of the main events governing start codon selection in eukaryotes and archaea occur within a common structural and functional core. However, idiosyncratic features in loop 1 sequence involved in 30S:mRNA binding suggest adjustments of e/AIF1 functioning in the two domains.
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Identification of a second GTP-bound magnesium ion in archaeal initiation factor 2
Nucleic Acids Research, 2016Co-Authors: Etienne Dubiez, Christine Lazennec-Schurdevin, Alexey Aleksandrov, Yves Mechulam, Emmanuelle SchmittAbstract:Eukaryotic and archaeal translation initiation processes involve a heterotrimeric GTPase e/aIF2 crucial for accuracy of start codon selection. In eu-karyotes, the GTPase activity of eIF2 is assisted by a GTPase-activating protein (GAP), eIF5. In ar-chaea, orthologs of eIF5 are not found and aIF2 GT-Pase activity is thought to be non-assisted. However , no in vitro GTPase activity of the archaeal factor has been reported to date. Here, we show that aIF2 significantly hydrolyses GTP in vitro. Within aIF2␥, H97, corresponding to the catalytic histidine found in other translational GTPases, and D19, from the GKT loop, both participate in this activity. Several high-resolution crystal structures were determined to get insight into GTP hydrolysis by aIF2␥. In particular, a crystal structure of the H97A mutant was obtained in the presence of non-hydrolyzed GTP. This structure reveals the presence of a second magnesium ion bound to GTP and D19. Quantum chemical/molecular mechanical simulations support the idea that the second magnesium ion may assist GTP hydrolysis by helping to neutralize the developing negative charge in the transition state. These results are discussed in light of the absence of an identified GAP in archaea to assist GTP hydrolysis on aIF2.
Udo Bläsi - One of the best experts on this subject based on the ideXlab platform.
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Translation initiation complex formation in the crenarchaeon Sulfolobus solfataricus
RNA (New York N.Y.), 2009Co-Authors: David Hasenohrl, Paola Londei, Attilio Fabbretti, Udo BläsiAbstract:The function of initiation factors in and the sequence of events during translation initiation have been intensively studied in Bacteria and Eukaryotes, whereas in Archaea knowledge on these functions/processes is limited. By employing chemical probing, we show that translation initiation factor AIF1 of the model crenarchaeon Sulfolobus solfataricus binds to the same area on the ribosome as the bacterial and eukaryal orthologs. Fluorescence energy transfer assays (FRET) showed that AIF1, like its eukaryotic and bacterial orthologs, has a fidelity function in translation initiation complex formation, and that both AIF1 and AIF1A exert a synergistic effect in stimulating ribosomal association of the Met-tRNAiMet binding factor a/eIF2. However, as in Eukaryotes their effect on a/eIF2 binding appears to be indirect. Moreover, FRET was used to analyze for the first time the sequence of events toward translation initiation complex formation in an archaeal model system. These studies suggested that a/eIF2-GTP binds first to the ribosome and then recruits Met-tRNAiMet, which appears to comply with the operational mode of bacterial IF2, and deviates from the shuttle function of the eukaryotic counterpart eIF2. Thus, despite the resemblance of eIF2 and a/eIF2, recruitment of initiator tRNA to the ribosome is mechanistically different in Pro- and Eukaryotes.
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sulfolobus solfataricus translation initiation factor 1 stimulates translation initiation complex formation
RNA, 2006Co-Authors: David Hasenohrl, Dario Benelli, A Barbazza, Paola Londei, Udo BläsiAbstract:The eukaryotic translation initiation factor 1 binds to the ribosome during translation initiation. It is instrumental for initiator-tRNA and mRNA binding, and has a function in selection of the authentic start codon. Here, we show that the archaeal homolog AIF1 has analogous functions. The AIF1 protein of the archaeon Sulfolobus solfataricus is bound to the small ribosomal subunit during translation initiation and accelerates binding of initiator-tRNA and mRNA to the ribosome. Accordingly, AIF1 stimulated translation of an mRNA in a S. solfataricus in vitro translation system. Moreover, this study suggested that the C terminus of the factor is of relevance for its function.
Pierre-damien Coureux - One of the best experts on this subject based on the ideXlab platform.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation
Communications Biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 Å resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life.
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Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation.
Communications biology, 2020Co-Authors: Pierre-damien Coureux, Christine Lazennec-Schurdevin, Yves Mechulam, Sophie Bourcier, Emmanuelle SchmittAbstract:Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors AIF1, AIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:AIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 A resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without AIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life. Coureux et al. describe a cryo-EM structure of a 30S initiation complex of Pyrococcus abyssi at 3.2A resolution. The structure uncovers a novel archaeal ribosomal protein aS21, N-terminal extension of eL41 and brings insights into base modifications of the rRNA.
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Start Codon Recognition in Eukaryotic and Archaeal Translation Initiation: A Common Structural Core
International Journal of Molecular Sciences, 2019Co-Authors: Emmanuelle Schmitt, Etienne Dubiez, Auriane Monestier, Pierre-damien Coureux, Yves MechulamAbstract:Understanding molecular mechanisms of ribosomal translation sheds light on the emergence and evolution of protein synthesis in the three domains of life. Universally, ribosomal translation is described in three steps: initiation, elongation and termination. During initiation, a macromolecular complex assembled around the small ribosomal subunit selects the start codon on the mRNA and defines the open reading frame. In this review, we focus on the comparison of start codon selection mechanisms in eukaryotes and archaea. Eukaryotic translation initiation is a very complicated process, involving many initiation factors. The most widespread mechanism for the discovery of the start codon is the scanning of the mRNA by a pre-initiation complex until the first AUG codon in a correct context is found. In archaea, long-range scanning does not occur because of the presence of Shine-Dalgarno (SD) sequences or of short 5' untranslated regions. However, archaeal and eukaryotic translation initiations have three initiation factors in common: e/AIF1, e/AIF1A and e/aIF2 are directly involved in the selection of the start codon. Therefore, the idea that these archaeal and eukaryotic factors fulfill similar functions within a common structural ribosomal core complex has emerged. A divergence between eukaryotic and archaeal factors allowed for the adaptation to the long-range scanning process versus the SD mediated prepositioning of the ribosome.
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Role of AIF1 in Pyrococcus abyssi translation initiation
Nucleic Acids Research, 2018Co-Authors: Auriane Monestier, Christine Lazennec-Schurdevin, Yves Mechulam, Pierre-damien Coureux, Emmanuelle SchmittAbstract:In archaeal translation initiation, a preinitiation complex (PIC) made up of AIF1, AIF1A, the ternary complex (TC, e/aIF2-GTP-Met-tRNA i Met) and mRNA bound to the small ribosomal subunit is responsible for start codon selection. Many archaeal mRNAs contain a Shine-Dalgarno (SD) sequence allowing the PIC to be prepositioned in the vicinity of the start codon. Nevertheless, cryo-EM studies have suggested local scanning to definitely establish base pairing of the start codon with the tRNA anticodon. Here, using flu-orescence anisotropy, we show that AIF1 and mRNA have synergistic binding to the Pyrococcus abyssi 30S. Stability of 30S:mRNA:AIF1 strongly depends on the SD sequence. Further, toeprinting experiments show that AIF1-containing PICs display a dynamic conformation with the tRNA not firmly accommodated in the P site. AIF1-induced destabilization of the PIC is favorable for proofreading erroneous initiation complexes. After AIF1 departure, the stability of the PIC increases reflecting initiator tRNA fully base-paired to the start codon. Altogether, our data support the idea that some of the main events governing start codon selection in eukaryotes and archaea occur within a common structural and functional core. However, idiosyncratic features in loop 1 sequence involved in 30S:mRNA binding suggest adjustments of e/AIF1 functioning in the two domains.
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Structure of the ternary initiation complex aIF2-GDPNP-methionylated initiator tRNA.
Nature Structural and Molecular Biology, 2012Co-Authors: Emmanuelle Schmitt, Christine Lazennec-Schurdevin, Pierre-damien Coureux, Michel Panvert, J. Perez, A. Thompson, Yves MechulamAbstract:Eukaryotic and archaeal translation initiation factor 2 (e/aIF2) is a heterotrimeric GTPase that has a crucial role in the selection of the correct start codon on messenger RNA. We report the 5-Å resolution crystal structure of the ternary complex formed by archaeal aIF2 from Sulfolobus solfataricus, the GTP analog GDPNP and methionylated initiator tRNA. The 3D model is further supported by solution studies using small-angle X-ray scattering. The tRNA is bound by the α and γ subunits of aIF2. Contacts involve the elbow of the tRNA and the minor groove of the acceptor stem, but not the T-stem minor groove. We conclude that despite considerable structural homology between the core γ subunit of aIF2 and the elongation factor EF1A, these two G proteins of the translation apparatus use very different tRNA-binding strategies.
David Hasenohrl - One of the best experts on this subject based on the ideXlab platform.
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Translation initiation complex formation in the crenarchaeon Sulfolobus solfataricus
RNA (New York N.Y.), 2009Co-Authors: David Hasenohrl, Paola Londei, Attilio Fabbretti, Udo BläsiAbstract:The function of initiation factors in and the sequence of events during translation initiation have been intensively studied in Bacteria and Eukaryotes, whereas in Archaea knowledge on these functions/processes is limited. By employing chemical probing, we show that translation initiation factor AIF1 of the model crenarchaeon Sulfolobus solfataricus binds to the same area on the ribosome as the bacterial and eukaryal orthologs. Fluorescence energy transfer assays (FRET) showed that AIF1, like its eukaryotic and bacterial orthologs, has a fidelity function in translation initiation complex formation, and that both AIF1 and AIF1A exert a synergistic effect in stimulating ribosomal association of the Met-tRNAiMet binding factor a/eIF2. However, as in Eukaryotes their effect on a/eIF2 binding appears to be indirect. Moreover, FRET was used to analyze for the first time the sequence of events toward translation initiation complex formation in an archaeal model system. These studies suggested that a/eIF2-GTP binds first to the ribosome and then recruits Met-tRNAiMet, which appears to comply with the operational mode of bacterial IF2, and deviates from the shuttle function of the eukaryotic counterpart eIF2. Thus, despite the resemblance of eIF2 and a/eIF2, recruitment of initiator tRNA to the ribosome is mechanistically different in Pro- and Eukaryotes.
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sulfolobus solfataricus translation initiation factor 1 stimulates translation initiation complex formation
RNA, 2006Co-Authors: David Hasenohrl, Dario Benelli, A Barbazza, Paola Londei, Udo BläsiAbstract:The eukaryotic translation initiation factor 1 binds to the ribosome during translation initiation. It is instrumental for initiator-tRNA and mRNA binding, and has a function in selection of the authentic start codon. Here, we show that the archaeal homolog AIF1 has analogous functions. The AIF1 protein of the archaeon Sulfolobus solfataricus is bound to the small ribosomal subunit during translation initiation and accelerates binding of initiator-tRNA and mRNA to the ribosome. Accordingly, AIF1 stimulated translation of an mRNA in a S. solfataricus in vitro translation system. Moreover, this study suggested that the C terminus of the factor is of relevance for its function.
