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Oxidation of cellular amino acid pools leads to cytotoxic mistranslation of the genetic code.

Bullwinkle TJ, Reynolds NM, Raina M, Moghal A, Matsa E, Rajkovic A, Kayadibi H, Fazlollahi F, Ryan C, Howitz N, Faull KF, Lazazzera BA, Ibba M - Elife (2014)

Bottom Line: Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis.The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict.These findings demonstrate how stress can lead to the accumulation of non-canonical amino acids that must be excluded from the proteome in order to maintain cellular viability.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Ohio State University, Columbus, United States.

ABSTRACT
Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. We show that in Escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein synthesis and must be cleared by phenylalanine-tRNA synthetase in order to prevent cellular toxicity caused by mis-synthesized proteins. These findings demonstrate how stress can lead to the accumulation of non-canonical amino acids that must be excluded from the proteome in order to maintain cellular viability.

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Related in: MedlinePlus

E. coli TyrRS uses m-Tyr.Aminoacylation of E. coli [32P]-tRNAPhe transcript (0.5 μM) with m-Tyr (1 mM) by E. coli TyrRS (50 nM) at 25°C.DOI:http://dx.doi.org/10.7554/eLife.02501.014
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fig6s2: E. coli TyrRS uses m-Tyr.Aminoacylation of E. coli [32P]-tRNAPhe transcript (0.5 μM) with m-Tyr (1 mM) by E. coli TyrRS (50 nM) at 25°C.DOI:http://dx.doi.org/10.7554/eLife.02501.014

Mentions: A detectable level of m-Tyr in the proteome of wild type E. coli suggests either this non-proteinogenic amino acid escapes PheRS editing, infiltrates the proteome by means other than misincorporation at Phe codons or is carried over during cytosolic protein preparation. To measure the approximate amount of carryover, wild type PheRS E. coli strain was grown in the presence of 0.5 mM o-Tyr, which is not a substrate for protein synthesis, and total protein samples were subjected to acid hydrolysis and LC-MS/MS-MRM. In these samples, traces of o-Tyr were detected, indicating that free amino acid carry over possibly contributes to some of the m-Tyr detected in the samples from the wild type strain grown in M9 minimal media supplemented with m-Tyr. Whether the m-Tyr seen in the proteome of E. coli containing PheRS editing is formed post-translationally or is incorporated during protein synthesis via another promiscuous tRNA synthetase in E. coli is unclear. Aminoacylation of tRNATyr with m-Tyr by E. coli TyrRS was detected in vitro, suggesting this synthetase may provide a route of m-Tyr incorporation even when PheRS editing is active (Figure 6—figure supplement 2).


Oxidation of cellular amino acid pools leads to cytotoxic mistranslation of the genetic code.

Bullwinkle TJ, Reynolds NM, Raina M, Moghal A, Matsa E, Rajkovic A, Kayadibi H, Fazlollahi F, Ryan C, Howitz N, Faull KF, Lazazzera BA, Ibba M - Elife (2014)

E. coli TyrRS uses m-Tyr.Aminoacylation of E. coli [32P]-tRNAPhe transcript (0.5 μM) with m-Tyr (1 mM) by E. coli TyrRS (50 nM) at 25°C.DOI:http://dx.doi.org/10.7554/eLife.02501.014
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4066437&req=5

fig6s2: E. coli TyrRS uses m-Tyr.Aminoacylation of E. coli [32P]-tRNAPhe transcript (0.5 μM) with m-Tyr (1 mM) by E. coli TyrRS (50 nM) at 25°C.DOI:http://dx.doi.org/10.7554/eLife.02501.014
Mentions: A detectable level of m-Tyr in the proteome of wild type E. coli suggests either this non-proteinogenic amino acid escapes PheRS editing, infiltrates the proteome by means other than misincorporation at Phe codons or is carried over during cytosolic protein preparation. To measure the approximate amount of carryover, wild type PheRS E. coli strain was grown in the presence of 0.5 mM o-Tyr, which is not a substrate for protein synthesis, and total protein samples were subjected to acid hydrolysis and LC-MS/MS-MRM. In these samples, traces of o-Tyr were detected, indicating that free amino acid carry over possibly contributes to some of the m-Tyr detected in the samples from the wild type strain grown in M9 minimal media supplemented with m-Tyr. Whether the m-Tyr seen in the proteome of E. coli containing PheRS editing is formed post-translationally or is incorporated during protein synthesis via another promiscuous tRNA synthetase in E. coli is unclear. Aminoacylation of tRNATyr with m-Tyr by E. coli TyrRS was detected in vitro, suggesting this synthetase may provide a route of m-Tyr incorporation even when PheRS editing is active (Figure 6—figure supplement 2).

Bottom Line: Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis.The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict.These findings demonstrate how stress can lead to the accumulation of non-canonical amino acids that must be excluded from the proteome in order to maintain cellular viability.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Ohio State University, Columbus, United States.

ABSTRACT
Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. We show that in Escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein synthesis and must be cleared by phenylalanine-tRNA synthetase in order to prevent cellular toxicity caused by mis-synthesized proteins. These findings demonstrate how stress can lead to the accumulation of non-canonical amino acids that must be excluded from the proteome in order to maintain cellular viability.

Show MeSH
Related in: MedlinePlus