Elongation Factor Tu Switch I Element is a Gate for Aminoacyl-tRNA Selection

Girodat, Dylan; Blanchard, Scott C.; Wieden, Hans-Joachim; Sanbonmatsu, Karissa Y.

doi:10.1016/j.jmb.2020.01.038

Cited by 12 publications

(7 citation statements)

References 78 publications

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“…All atom Gō-like structure-based simulations were performed as previously described [ 48 , 66 ]. AMBER minimized structures of the apo LIV-BP SS protein or bound to Leu, Ile, or Val (excluding hydrogens) were used as starting structures for structure-based models.…”

Section: Methodsmentioning

confidence: 99%

“…For this technique, specific hydrogen bonds, electrostatic interactions, and rotamer angles are implicitly included in the native basin. By combining this approach with explicit-solvent equilibrium simulations, one can achieve comprehensive views of the dynamic ensemble of a biomolecule by explicitly resolving even more subtle interactions, such as explicit hydrogen bonds, electrostatics, or solvation effects [ 65 , 66 ].…”

Section: Introductionmentioning

confidence: 99%

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Quantitative comparison between sub-millisecond time resolution single-molecule FRET measurements and 10-second molecular simulations of a biosensor protein

et al. 2020

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Molecular Dynamics (MD) simulations seek to provide atomic-level insights into conformationally dynamic biological systems at experimentally relevant time resolutions, such as those afforded by single-molecule fluorescence measurements. However, limitations in the time scales of MD simulations and the time resolution of single-molecule measurements have challenged efforts to obtain overlapping temporal regimes required for close quantitative comparisons. Achieving such overlap has the potential to provide novel theories, hypotheses, and interpretations that can inform idealized experimental designs that maximize the detection of the desired reaction coordinate. Here, we report MD simulations at time scales overlapping with in vitro single-molecule Förster (fluorescence) resonance energy transfer (smFRET) measurements of the amino acid binding protein LIV-BPSS at sub-millisecond resolution. Computationally efficient all-atom structure-based simulations, calibrated against explicit solvent simulations, were employed for sampling multiple cycles of LIV-BPSS clamshell-like conformational changes on the time scale of seconds, examining the relationship between these events and those observed by smFRET. The MD simulations agree with the smFRET measurements and provide valuable information on local dynamics of fluorophores at their sites of attachment on LIV-BPSS and the correlations between fluorophore motions and large-scale conformational changes between LIV-BPSS domains. We further utilize the MD simulations to inform the interpretation of smFRET data, including Förster radius (R0) and fluorophore orientation factor (κ2) determinations. The approach we describe can be readily extended to distinct biochemical systems, allowing for the interpretation of any FRET system conjugated to protein or ribonucleoprotein complexes, including those with more conformational processes, as well as those implementing multi-color smFRET.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Quantitative comparison between sub-millisecond time resolution single-molecule FRET measurements and 10-second molecular simulations of a biosensor protein

et al. 2020

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“…This includes EF-Tu, a translational GTPase, which transports aminoacylated tRNA to the ribosomal A-site and regulates tRNA selection and proofreading. 61 As the switch I region of EF-Tu has been proposed to interact with the acceptor stem of tRNA 62 during proofreading, tRNA CUD likely does not affect the fidelity mechanism of EF-Tu. Remarkably, the ribosome could tolerate ASL-modified tRNA CUD during translation.…”

Section: ■ Conclusionmentioning

confidence: 99%

Enhanced Codon–Anticodon Interaction at In-Frame UAG Stop Codon Improves the Efficiency of Non-Natural Amino Acid Mutagenesis

Saraogi

2022

ACS Chem. Biol.

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The introduction of non-natural amino acids into proteins through the stop codon readthrough methodology has been used to design proteins for diverse applications. However, this method suffers from low yields of the modified protein, as the suppressor tRNA that recognizes the stop codon is unable to compete effectively with release factor 1 (RF1), which terminates translation. We reasoned that a suppressor tRNA with improved interaction with the UAG stop codon on the mRNA will be able to compete more effectively with RF1. To test this idea, we inserted two 2,6-diaminopurine (D) units in the tRNA anticodon stem loop, including one at the third position of the tRNA anticodon. The modified suppressor tRNA could potentially form additional H-bonds between the N 2 -exocyclic amine of D and the C2 carbonyl group of uracil, thereby enhancing mRNA−tRNA interaction and/or altering tRNA conformation. The stronger interaction at the codon−anticodon interface resulted in improved UAG decoding efficiency and a higher yield of the modified protein containing a non-natural amino acid at multiple sites. Our findings are consistent with the importance of hydrogen bonding and tRNA conformation at the tRNA−mRNA duplex interface during in-frame UAG suppression, which improves protein translation at multiple UAG stop sites. This work provides valuable inputs toward improved non-natural amino acid mutagenesis for creating designer proteins.

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“…It is an important step for EFTU to select the correct aa-tRNA for the ribosome A site to ensure the fidelity of translation. Using structure-based and explicit solvent molecular dynamics simulations based on recent cryo-EM reconstructions, Girodat et al (2020) investigated the structural mechanism of how EFTU is involved in proofreading. They found that switch I of EFTU is a gate that facilitates aa-tRNA selection.…”

Section: Mitochondrial Translation Elongation and Diseasementioning

confidence: 99%

Mitochondrial Protein Translation: Emerging Roles and Clinical Significance in Disease

Wang

Zhang

et al. 2021

Front. Cell Dev. Biol.

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Mitochondria are one of the most important organelles in cells. Mitochondria are semi-autonomous organelles with their own genetic system, and can independently replicate, transcribe, and translate mitochondrial DNA. Translation initiation, elongation, termination, and recycling of the ribosome are four stages in the process of mitochondrial protein translation. In this process, mitochondrial protein translation factors and translation activators, mitochondrial RNA, and other regulatory factors regulate mitochondrial protein translation. Mitochondrial protein translation abnormalities are associated with a variety of diseases, including cancer, cardiovascular diseases, and nervous system diseases. Mutation or deletion of various mitochondrial protein translation factors and translation activators leads to abnormal mitochondrial protein translation. Mitochondrial tRNAs and mitochondrial ribosomal proteins are essential players during translation and mutations in genes encoding them represent a large fraction of mitochondrial diseases. Moreover, there is crosstalk between mitochondrial protein translation and cytoplasmic translation, and the imbalance between mitochondrial protein translation and cytoplasmic translation can affect some physiological and pathological processes. This review summarizes the regulation of mitochondrial protein translation factors, mitochondrial ribosomal proteins, mitochondrial tRNAs, and mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) in the mitochondrial protein translation process and its relationship with diseases. The regulation of mitochondrial protein translation and cytoplasmic translation in multiple diseases is also summarized.

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Elongation Factor Tu Switch I Element is a Gate for Aminoacyl-tRNA Selection

Cited by 12 publications

References 78 publications

Quantitative comparison between sub-millisecond time resolution single-molecule FRET measurements and 10-second molecular simulations of a biosensor protein

Quantitative comparison between sub-millisecond time resolution single-molecule FRET measurements and 10-second molecular simulations of a biosensor protein

Enhanced Codon–Anticodon Interaction at In-Frame UAG Stop Codon Improves the Efficiency of Non-Natural Amino Acid Mutagenesis

Mitochondrial Protein Translation: Emerging Roles and Clinical Significance in Disease

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