Elongation Factor Ts Directly Facilitates the Formation and Disassembly of the Escherichia coli Elongation Factor Tu·GTP·Aminoacyl-tRNA Ternary Complex

Burnett, Benjamin J.; Altman, Roger B.; Ferrao, Ryan; Alejo, José Luis Pérez; Kaur, Navdep; Kanji, Joshua; Blanchard, Scott C.

doi:10.1074/jbc.m113.460014

Cited by 38 publications

(79 citation statements)

References 67 publications

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“…We propose that EF-Ts acts as a critical accessory factor to load tRNA onto EF-Tu, accelerating the production of GTP·EF-Tu·tRNA substrate for cleavage. This model is supported by recent work from Blanchard and colleagues, who have shown that EF-Ts promotes both the association and dissociation of tRNA from EF-Tu (29). This activity is concomitant to yet distinct from the well-described GEF function of EF-Ts (30).…”

Section: Discussionsupporting

confidence: 68%

Activation of contact-dependent antibacterial tRNase toxins by translation elongation factors

Jones

Garza-Sánchez

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Contact-dependent growth inhibition (CDI) is a mechanism by which bacteria exchange toxins via direct cell-to-cell contact. CDI systems are distributed widely among Gram-negative pathogens and are thought to mediate interstrain competition. Here, we describe tsf mutations that alter the coiled-coil domain of elongation factor Ts (EF-Ts) and confer resistance to the CdiA-CT EC869 tRNase toxin from enterohemorrhagic Escherichia coli EC869. Although EF-Ts is required for toxicity in vivo, our results indicate that it is dispensable for tRNase activity in vitro. We find that CdiA-CT EC869 binds to elongation factor Tu (EF-Tu) with high affinity and this interaction is critical for nuclease activity. Moreover, in vitro tRNase activity is GTP-dependent, suggesting that CdiA-CT EC869 only cleaves tRNA in the context of translationally active GTP·EF-Tu·tRNA ternary complexes. We propose that EF-Ts promotes the formation of GTP·EF-Tu·tRNA ternary complexes, thereby accelerating substrate turnover for rapid depletion of target-cell tRNA.cell communication | protein synthesis | toxin-immunity proteins | ribonuclease | bacterial competition

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Section: Discussionsupporting

confidence: 68%

Activation of contact-dependent antibacterial tRNase toxins by translation elongation factors

Jones

Garza-Sánchez

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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“…After removal of His 6 , the elongation factors were purified further using Superdex 75 gel filtration chromatography as described previously (1).…”

Section: Methodsmentioning

confidence: 99%

“…These observations suggested that EF-Ts has a capacity to directly regulate ternary complex concentration via the formation of an EF-Tu/Ts⅐GTP⅐aa-tRNA quaternary complex (1). Here, we directly examined whether such findings are general in nature by measuring the impact of EF-Ts on the kinetic properties of distinct ternary complex species that exhibit distinct apparent K D values (29).…”

mentioning

confidence: 99%

Direct Evidence of an Elongation Factor-Tu/Ts·GTP·Aminoacyl-tRNA Quaternary Complex

Burnett

Altman

Ferguson

et al. 2014

Journal of Biological Chemistry

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Background: Elongation factor-Tu (EF-Tu) chaperones aminoacyl-tRNA (aa-tRNA) to the elongating ribosome as a ternary complex with GTP. Results: EF-Ts facilitates EF-Tu⅐GTP binding to aa-tRNA through direct, transient interactions. Conclusion: EF-Ts accelerates the formation and decay of ternary complex through the formation of a transient EF-Tu/ Ts⅐GTP⅐aa-tRNA quaternary complex. Significance: These newly described interactions of EF-Ts may serve to regulate ternary complex abundance in the cell.

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“…1B). Based on high cellular concentrations of EF-Tu (100-200 μM) (Burnett et al 2013) and the entire population of aa-tRNAs (50-200 μM) (Dong et al 1996), the protection of EF-Tu-GTP at a high molar ratio to a single Pro-tRNA Pro species is physiologically relevant. This protection is consistent with an earlier observation of EF-Tu-GTP protecting the acyl stability of Phe-tRNA Phe by more than 10-fold (from T 1/2 of 56 to 800 min) (Beres and Lucas-Lenard 1973).…”

Section: Analysis Of Amino Acid Enantiomerism and Ef-tu Effects On Stmentioning

confidence: 99%

Amino acid–dependent stability of the acyl linkage in aminoacyl-tRNA

Peacock

Walvoord

et al. 2014

RNA

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Aminoacyl-tRNAs are the biologically active substrates for peptide bond formation in protein synthesis. The stability of the acyl linkage in each aminoacyl-tRNA, formed through an ester bond that connects the amino acid carboxyl group with the tRNA terminal 3′ -OH group, is thus important. While the ester linkage is the same for all aminoacyl-tRNAs, the stability of each is not well characterized, thus limiting insight into the fundamental process of peptide bond formation. Here, we show, by analysis of the half-lives of 12 of the 22 natural aminoacyl-tRNAs used in peptide bond formation, that the stability of the acyl linkage is effectively determined only by the chemical nature of the amino acid side chain. Even the chirality of the side chain exhibits little influence. Proline confers the lowest stability to the linkage, while isoleucine and valine confer the highest, whereas the nucleotide sequence in the tRNA provides negligible contribution to the stability. We find that, among the variables tested, the protein translation factor EF-Tu is the only one that can protect a weak acyl linkage from hydrolysis. These results suggest that each amino acid plays an active role in determining its own stability in the acyl linkage to tRNA, but that EF-Tu overrides this individuality and protects the acyl linkage stability for protein synthesis on the ribosome.

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Elongation Factor Ts Directly Facilitates the Formation and Disassembly of the Escherichia coli Elongation Factor Tu·GTP·Aminoacyl-tRNA Ternary Complex

Cited by 38 publications

References 67 publications

Activation of contact-dependent antibacterial tRNase toxins by translation elongation factors

Activation of contact-dependent antibacterial tRNase toxins by translation elongation factors

Direct Evidence of an Elongation Factor-Tu/Ts·GTP·Aminoacyl-tRNA Quaternary Complex

Amino acid–dependent stability of the acyl linkage in aminoacyl-tRNA

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