The pathway to GTPase activation of elongation factor SelB on the ribosome

Fischer, N.; Neumann, Piotr; Bock, Lars V.; Maracci, Cristina; Wang, Zhe; Paleskava, Alena; Konevega, Andrey L.; Schröder, Gunnar F.; Grubmüller, Helmut; Ficner, Ralf; Rodnina, Marina V.; Stark, Holger

doi:10.1038/nature20560

Cited by 106 publications

(174 citation statements)

References 95 publications

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“…The molecular model for the ribosomal proteins and rRNA of the 70S ribosome of the VemP-SRC was based on the molecular model from cryo-EM reconstruction of the E. coli 70S ribosome (PDB ID 5JU8) (Arenz et al, 2016), except that the bL31 was based on the (PDB ID 5LZD) (Fischer et al, 2016). The molecular model for Gln-tRNA was based on a crystal structure (PDB ID 1GSG) (Rould et al, 1989).…”

Section: Methodsmentioning

confidence: 99%

The force-sensing peptide VemP employs extreme compaction and secondary structure formation to induce ribosomal stalling

Cheng

Sohmen

et al. 2017

eLife

113

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Interaction between the nascent polypeptide chain and the ribosomal exit tunnel can modulate the rate of translation and induce translational arrest to regulate expression of downstream genes. The ribosomal tunnel also provides a protected environment for initial protein folding events. Here, we present a 2.9 Å cryo-electron microscopy structure of a ribosome stalled during translation of the extremely compacted VemP nascent chain. The nascent chain forms two α-helices connected by an α-turn and a loop, enabling a total of 37 amino acids to be observed within the first 50–55 Å of the exit tunnel. The structure reveals how α-helix formation directly within the peptidyltransferase center of the ribosome interferes with aminoacyl-tRNA accommodation, suggesting that during canonical translation, a major role of the exit tunnel is to prevent excessive secondary structure formation that can interfere with the peptidyltransferase activity of the ribosome.DOI: http://dx.doi.org/10.7554/eLife.25642.001

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Section: Methodsmentioning

confidence: 99%

The force-sensing peptide VemP employs extreme compaction and secondary structure formation to induce ribosomal stalling

Cheng

Sohmen

et al. 2017

eLife

113

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“…It has been proposed that the ALC represents the GAC rearrangement and may play an important role for simulation of GTP hydrolysis by trGTPases. The mechanism of GTP hydrolysis is conserved in all domains of life (35,36), with the SRL being critical for direct activation of the trGTPases (37,38). However, the cooperativity of the SRL and both the base and lateral stalk elements has been shown to be pivotal in the stimulation of GTP hydrolysis by trGTPases (39,40).…”

mentioning

confidence: 99%

Multiplication of Ribosomal P-Stalk Proteins Contributes to the Fidelity of Translation

Wawiórka

Molestak

Szajwaj

et al. 2017

Molecular and Cellular Biology

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The P-stalk represents a vital element within the ribosomal GTPaseassociated center, which represents a landing platform for translational GTPases. The eukaryotic P-stalk exists as a uL10-(P1-P2) 2 pentameric complex, which contains five identical C-terminal domains, one within each protein, and the presence of only one such element is sufficient to stimulate factor-dependent GTP hydrolysis in vitro and to sustain cell viability. The functional contribution of the P-stalk to the performance of the translational machinery in vivo, especially the role of P-protein multiplication, has never been explored. Here, we show that ribosomes depleted of P1/P2 proteins exhibit reduced translation fidelity at elongation and termination steps. The elevated rate of the decoding error is inversely correlated with the number of the P-proteins present on the ribosome. Unexpectedly, the lack of P1/P2 has little effect in vivo on the efficiency of other translational GTPase (trGTPase)-dependent steps of protein synthesis, including translocation. We have shown that loss of accuracy of decoding caused by P1/P2 depletion is the major cause of translation slowdown, which in turn affects the metabolic fitness of the yeast cell. We postulate that the multiplication of P-proteins is functionally coupled with the qualitative aspect of ribosome action, i.e., the recoding phenomenon shaping the cellular proteome.KEYWORDS ribosomal proteins, ribosomal stalk, ribosome A t the expense of energy from GTP hydrolysis, translational GTPases (trGTPases) confer the unidirectional trajectory for the translational apparatus, providing at the same time unique timing for individual steps (1, 2). The main landing platform for trGTPases is situated on the large ribosomal subunit called the GTPase-associated center (GAC), and it represents a universally conserved ribosomal element where stimulation of trGTPase catalytic activity takes place (3). The GAC consists of two main elements, a conserved fragment of rRNA called the sarcin-ricin loop (SRL) and a ribosomal stalk composed of ribosomal proteins, which form an oligomeric protein complex (4). The protein part of GAC, the ribosome stalk, can be divided into two functionally and evolutionarily distinct parts, the base of the stalk and its lateral elements. The stalk base is composed of conserved ribosomal proteins uL11 (former names L11 and L12 for prokaryotes and eukaryotes, respectively) and uL10 (former names L10 and P0), which anchor the stalk to the rRNA (5, 6). The lateral part of the stalk is built of dimeric complexes P1-P2 in eukaryotes/archaea or (bL12) 2 in prokaryotes (4, 7). Despite the lack of amino acid sequence conservation, the lateral stalk fulfils the same functions and has a similar structural architecture across all domains of life (8). P1/P2 and bL12 proteins are built of two domains. The globular N-terminal domain (NTD) is responsible for dimerization, whereas the highly acidic C-terminal domain (CTD) interacts with trGTPases (9, 10). However, the structure of the CTD in euk...

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“…The genetic code of mitochondria deviates significantly from the standard genetic code (61, 76) (Supplemental Figure 1), probably because there are only a few genes (encoding membrane proteins) in mitochondrial genomes. It was also known that the UGA stop codon is recoded as selenocysteine (Sec), the twenty-first genetically encoded amino acid, in the three domains of life when guided by Sec-insertion sequence elements (7, 33, 93, 158). In contrast, the UAG stop codon is translated as pyrrolysine (Pyl), the twenty-second genetically encoded amino acid, in some anaerobic archaea and bacteria (35, 93, 158), and as glutamine (Gln) in the late genes of some bacteriophages that kill bacteria whose UGA codon encodes Trp (54) (Figure 2).…”

Section: Natural Expansion Of the Genetic Codementioning

confidence: 99%

“…Although Sec naturally requires the dedicated elongation factor SelB (33), two separate tRNAs have been reengineered to incorporate Sec using the more conventional elongation factor EF-Tu (2, 44, 89, 139), which is not dependent on a Sec insertion sequence in the mRNA. Finally, it would be beneficial to develop orthogonal aaRS•tRNA pairs that can function across all domains, like the tRNA Pyl •PylRS pair (97).…”

Section: Expanding the Genetic Code With Orthogonal Translation Systemsmentioning

confidence: 99%

Rewriting the Genetic Code

Mukai

Lajoie

Englert

et al. 2017

Annu. Rev. Microbiol.

145

137

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The genetic code—the language used by cells to translate their genomes into proteins that perform many cellular functions—is highly conserved throughout natural life. Rewriting the genetic code could lead to new biological functions such as expanding protein chemistries with noncanonical amino acids (ncAAs) and genetically isolating synthetic organisms from natural organisms and viruses. It has long been possible to transiently produce proteins bearing ncAAs, but stabilizing an expanded genetic code for sustained function in vivo requires an integrated approach: creating recoded genomes and introducing new translation machinery that function together without compromising viability or clashing with endogenous pathways. In this review, we discuss design considerations and technologies for expanding the genetic code. The knowledge obtained by rewriting the genetic code will deepen our understanding of how genomes are designed and how the canonical genetic code evolved.

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The pathway to GTPase activation of elongation factor SelB on the ribosome

Cited by 106 publications

References 95 publications

The force-sensing peptide VemP employs extreme compaction and secondary structure formation to induce ribosomal stalling

The force-sensing peptide VemP employs extreme compaction and secondary structure formation to induce ribosomal stalling

Multiplication of Ribosomal P-Stalk Proteins Contributes to the Fidelity of Translation

Rewriting the Genetic Code

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