Selenocysteine Inserting RNA Elements Modulate GTP Hydrolysis of Elongation Factor SelB†

Biochemistry ◽  
1998 ◽  
Vol 37 (3) ◽  
pp. 885-890 ◽  
Author(s):  
Alexander Hüttenhofer ◽  
August Böck
Keyword(s):  
Elongation Factor ◽  
Gtp Hydrolysis ◽  
Rna Elements ◽  
Hydrolysis Of

scholarly journals Ribosome dynamics during decoding

2017 ◽  
Vol 372 (1716) ◽  
pp. 20160182 ◽  
Author(s):  
Marina V. Rodnina ◽  
Niels Fischer ◽  
Cristina Maracci ◽  
Holger Stark
Keyword(s):  
Translation Termination ◽  
Elongation Factor ◽  
Translation Elongation ◽  
Gtp Hydrolysis ◽  
Codon Recognition ◽  
Special Cases ◽  
Multistep Process ◽  
Gtpase Activation ◽  
Hydrolysis Of

Elongation factors Tu (EF-Tu) and SelB are translational GTPases that deliver aminoacyl-tRNAs (aa-tRNAs) to the ribosome. In each canonical round of translation elongation, aa-tRNAs, assisted by EF-Tu, decode mRNA codons and insert the respective amino acid into the growing peptide chain. Stop codons usually lead to translation termination; however, in special cases UGA codons are recoded to selenocysteine (Sec) with the help of SelB. Recruitment of EF-Tu and SelB together with their respective aa-tRNAs to the ribosome is a multistep process. In this review, we summarize recent progress in understanding the role of ribosome dynamics in aa-tRNA selection. We describe the path to correct codon recognition by canonical elongator aa-tRNA and Sec-tRNA Sec and discuss the local and global rearrangements of the ribosome in response to correct and incorrect aa-tRNAs. We present the mechanisms of GTPase activation and GTP hydrolysis of EF-Tu and SelB and summarize what is known about the accommodation of aa-tRNA on the ribosome after its release from the elongation factor. We show how ribosome dynamics ensures high selectivity for the cognate aa-tRNA and suggest that conformational fluctuations, induced fit and kinetic discrimination play major roles in maintaining the speed and fidelity of translation. This article is part of the themed issue ‘Perspectives on the ribosome’.

scholarly journals RASAL3 preferentially stimulates GTP hydrolysis of the Rho family small GTPase Rac2

2018 ◽  
Author(s):  
Yoonjae Shin ◽  
Yong Kim ◽  
Hyemin Kim ◽  
Nakyoung Shin ◽  
Tae Kim ◽  
...  
Keyword(s):  
Small Gtpase ◽  
Gtp Hydrolysis ◽  
Rho Family ◽  
Hydrolysis Of

scholarly journals Role of a ribosomal RNA phosphate oxygen during the EF-G–triggered GTP hydrolysis

2015 ◽  
Vol 112 (20) ◽  
pp. E2561-E2568 ◽  
Author(s):  
Miriam Koch ◽  
Sara Flür ◽  
Christoph Kreutz ◽  
Eric Ennifar ◽  
Ronald Micura ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
Electrostatic Interactions ◽  
Elongation Factor ◽  
Direct Interaction ◽  
Phosphate Group ◽  
Gtp Hydrolysis ◽  
Elongation Cycle ◽  
Close Proximity ◽  
Catalytically Active ◽  
Gtpase Activation

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

The ribosome as a molecular machine: the mechanism of tRNA–mRNA movement in translocation

2011 ◽  
Vol 39 (2) ◽  
pp. 658-662 ◽  
Author(s):  
Marina V. Rodnina ◽  
Wolfgang Wintermeyer
Keyword(s):  
Ribosomal Subunit ◽  
Elongation Factor ◽  
Thermal Fluctuations ◽  
Spontaneous Movement ◽  
Conformational States ◽  
Gtp Hydrolysis ◽  
Time Resolved ◽  
Directional Bias ◽  
Loosely Coupled ◽  
Dynamic Events

Translocation of tRNA and mRNA through the ribosome is one of the most dynamic events during protein synthesis. In the cell, translocation is catalysed by EF-G (elongation factor G) and driven by GTP hydrolysis. Major unresolved questions are: how the movement is induced and what the moving parts of the ribosome are. Recent progress in time-resolved cryoelectron microscopy revealed trajectories of tRNA movement through the ribosome. Driven by thermal fluctuations, the ribosome spontaneously samples a large number of conformational states. The spontaneous movement of tRNAs through the ribosome is loosely coupled to the motions within the ribosome. EF-G stabilizes conformational states prone to translocation and promotes a conformational rearrangement of the ribosome (unlocking) that accelerates the rate-limiting step of translocation: the movement of the tRNA anticodons on the small ribosomal subunit. EF-G acts as a Brownian ratchet providing directional bias for movement at the cost of GTP hydrolysis.

Binding of GDP to a ribosomal protein after elongation factor-2 dependent GTP hydrolysis

1992 ◽  
Vol 1132 (3) ◽  
pp. 284-289 ◽  
Author(s):  
Jean-Pierre Lavergne ◽  
Anne-Marie Reboud ◽  
Bruno Sontag ◽  
Dominique Guillot ◽  
Jean-Paul Reboud
Keyword(s):  
Ribosomal Protein ◽  
Elongation Factor ◽  
Gtp Hydrolysis ◽  
Elongation Factor 2

scholarly journals Coordination of the leucine-sensing Rag GTPase cycle by leucyl-tRNA synthetase in the mTORC1 signaling pathway

2018 ◽  
Vol 115 (23) ◽  
pp. E5279-E5288 ◽  
Author(s):  
Minji Lee ◽  
Jong Hyun Kim ◽  
Ina Yoon ◽  
Chulho Lee ◽  
Mohammad Fallahi Sichani ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Trna Synthetase ◽  
Gtp Hydrolysis ◽  
Mtorc1 Signaling ◽  
Amino Acid Sensing ◽  
Gtpase Cycle ◽  
Rag Gtpase ◽  
Cancer Tissues ◽  
Mtorc1 Activation ◽  
Hydrolysis Of

A protein synthesis enzyme, leucyl-tRNA synthetase (LRS), serves as a leucine sensor for the mechanistic target of rapamycin complex 1 (mTORC1), which is a central effector for protein synthesis, metabolism, autophagy, and cell growth. However, its significance in mTORC1 signaling and cancer growth and its functional relationship with other suggested leucine signal mediators are not well-understood. Here we show the kinetics of the Rag GTPase cycle during leucine signaling and that LRS serves as an initiating “ON” switch via GTP hydrolysis of RagD that drives the entire Rag GTPase cycle, whereas Sestrin2 functions as an “OFF” switch by controlling GTP hydrolysis of RagB in the Rag GTPase–mTORC1 axis. The LRS–RagD axis showed a positive correlation with mTORC1 activity in cancer tissues and cells. The GTP–GDP cycle of the RagD–RagB pair, rather than the RagC–RagA pair, is critical for leucine-induced mTORC1 activation. The active RagD–RagB pair can overcome the absence of the RagC–RagA pair, but the opposite is not the case. This work suggests that the GTPase cycle of RagD–RagB coordinated by LRS and Sestrin2 is critical for controlling mTORC1 activation, and thus will extend the current understanding of the amino acid-sensing mechanism.

scholarly journals EF-G–induced ribosome sliding along the noncoding mRNA

2019 ◽  
Vol 5 (6) ◽  
pp. eaaw9049 ◽  
Author(s):  
M. Klimova ◽  
T. Senyushkina ◽  
E. Samatova ◽  
B. Z. Peng ◽  
M. Pearson ◽  
...  
Keyword(s):  
Messenger Rna ◽  
Elongation Factor ◽  
Transfer Rna ◽  
Noncoding Region ◽  
Forward Movement ◽  
Stem Loop ◽  
A Site ◽  
Hydrolysis Of ◽  
Reading Frames ◽  
Factor G

Translational bypassing is a recoding event during which ribosomes slide over a noncoding region of the messenger RNA (mRNA) to synthesize one protein from two discontinuous reading frames. Structures in the mRNA orchestrate forward movement of the ribosome, but what causes ribosomes to start sliding remains unclear. Here, we show that elongation factor G (EF-G) triggers ribosome take-off by a pseudotranslocation event using a small mRNA stem-loop as an A-site transfer RNA mimic and requires hydrolysis of about two molecules of guanosine 5′-triphosphate per nucleotide of the noncoding gap. Bypassing ribosomes adopt a hyper-rotated conformation, also observed with ribosomes stalled by the SecM sequence, suggesting common ribosome dynamics during translation stalling. Our results demonstrate a new function of EF-G in promoting ribosome sliding along the mRNA, in contrast to codon-wise ribosome movement during canonical translation, and suggest a mechanism by which ribosomes could traverse untranslated parts of mRNAs.

scholarly journals Irreversible chemical steps control intersubunit dynamics during translation

2008 ◽  
Vol 105 (40) ◽  
pp. 15364-15369 ◽  
Author(s):  
R. Andrew Marshall ◽  
Magdalena Dorywalska ◽  
Joseph D. Puglisi
Keyword(s):  
Single Molecule ◽  
Conformational Changes ◽  
Large Scale ◽  
Bond Formation ◽  
Elongation Factor ◽  
Resonance Energy ◽  
Peptide Bond ◽  
Peptide Bond Formation ◽  
Gtp Hydrolysis ◽  
30S Subunit

The ribosome, a two-subunit macromolecular machine, deciphers the genetic code and catalyzes peptide bond formation. Dynamic rotational movement between ribosomal subunits is likely required for efficient and accurate protein synthesis, but direct observation of intersubunit dynamics has been obscured by the repetitive, multistep nature of translation. Here, we report a collection of single-molecule fluorescence resonance energy transfer assays that reveal a ribosomal intersubunit conformational cycle in real time during initiation and the first round of elongation. After subunit joining and delivery of correct aminoacyl-tRNA to the ribosome, peptide bond formation results in a rapid conformational change, consistent with the counterclockwise rotation of the 30S subunit with respect to the 50S subunit implied by prior structural and biochemical studies. Subsequent binding of elongation factor G and GTP hydrolysis results in a clockwise rotation of the 30S subunit relative to the 50S subunit, preparing the ribosome for the next round of tRNA selection and peptide bond formation. The ribosome thus harnesses the free energy of irreversible peptidyl transfer and GTP hydrolysis to surmount activation barriers to large-scale conformational changes during translation. Intersubunit rotation is likely a requirement for the concerted movement of tRNA and mRNA substrates during translocation.

scholarly journals Polarization of Diploid Daughter Cells Directed by Spatial Cues and GTP Hydrolysis of Cdc42 in Budding Yeast

PLoS ONE ◽  
2013 ◽  
Vol 8 (2) ◽  
pp. e56665 ◽  
Author(s):  
Wing-Cheong Lo ◽  
Mid Eum Lee ◽  
Monisha Narayan ◽  
Ching-Shan Chou ◽  
Hay-Oak Park
Keyword(s):  
Budding Yeast ◽  
Spatial Cues ◽  
Gtp Hydrolysis ◽  
Daughter Cells ◽  
Hydrolysis Of
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