Transfer RNA Synthetase Editing of Errors in Amino Acid Selection


Aminoacyl‐tRNA synthetases (AARSs) define genetic code by pairing amino acids with their cognate tRNAs and maintain ‘quality control’ in the flow of information from gene to protein. Binding energies of amino acids to AARSs are often inadequate to assure the required accuracy of translation. This has necessitated the evolution of a second determinant of specificity, proofreading or editing mechanisms that remove amino acid selection errors, thereby maintaining accuracy and preventing access to the genetic code of non‐protein amino acids, including homocysteine, ornithine, homoserine and norvaline. Editing is part of the tRNA aminoacylation process in living organisms from bacteria to humans. Impaired amino acid editing reduces cell proliferation under stress conditions and can lead to disease. Despite a strong selective pressure to minimise mistranslation, some organisms possess error‐prone AARSs that cause mistranslation, which can be beneficial for pathogens by increasing their phenotypic variation essential for the evasion of host defences.

Key Concepts

  • Aminoacyl‐tRNA synthetases match tRNAs with corresponding amino acids, thereby translating genetic information from the nucleic acid to the protein language.
  • Faithful translation of the genetic information depends on editing of amino acid misactivation errors.
  • Editing occurs at the synthetic active site by hydrolysis of aminoacyl‐adenylates (pre‐transfer editing) and/or at a separate editing site by deacylation of aminoacyl‐tRNA (post‐transfer editing).
  • Access of the non‐protein amino acids, such as homocysteine, ornithine or norvaline to the genetic code is prevented by editing.
  • Preventing mistranslation by editing of misactivated amino acids is crucial to cellular homeostasis.
  • Impaired editing increases sensitivity of cells to oxidative stress and nutritional imbalance.
  • Some pathogenic organisms possess editing‐defective, error‐prone AARSs that cause mistranslation, which is essential for the evasion of host defences.
  • Synthetic biology targets AARSs to engineer new proteins containing non‐canonical amino acids (alloproteins).

Keywords: genetic code; protein biosynthesis; amino acid selection and editing; mistranslation errors; cardiovascular disease risk factors; hyperhomocysteinaemia and neurodegeneration; protein engineering; synthetic biology; aminoacyl‐coenzyme A thioesters; prebiotic chemistry

Figure 1. Substrate‐assisted catalysis of hydrolytic deacylation of mischarged Ser‐tRNAThr in ThrRS editing domain. The 2′ OH of the ribose acts as a general base and activates water molecule (W1) for a nucleophilic attack on the carbonyl carbon of the L‐Ser‐tRNAThr. The tetrahedral intermediate is stabilised by the oxyanion hole formed by the main chain nitrogen atom of Ala82 and main chain as well as side‐chain nitrogen atoms of His83. Reproduced with permission from Hussain et al. (2010) © National Academy of Sciences .
Figure 3. Structural rearrangements in the active site of mitochondrial ThrRS (MST1) promoted by binding of a non‐hydrolysable Ser‐AMP analogue, seryl sulphamoyl adenylate (SAM). (a) Structural comparison between MST1•TAM ( ; PDB ID code 3UH0) and MST1•SAM ( ) reveals that a number of side chains and water molecules adopt a different orientation when SAM ( ) binds to the active site. The Tyr109 and Asp112 residues ( ) from helix α4′, and water molecules Wat1 and Wat2 ( ), are positioned closer to SAM than to threonyl sulphoamyl adenylate (TAM), a Thr‐AMP analogue ( ). Wat1, a putative hydrolytic water, is positioned differently in MST1•TAM; the water in that complex is designated as Wat1′ and is shown as a (b) hydrogen‐bonding network in the active site of MST1•SAM positioning the putative hydrolytic water (Wat1) at a distance and an angle proper for nucleophilic attack on the phosphorus atom mimic. All hydrogen bonds are shown as . Reproduced with permission from Ling et al. (2012). © The American Society for Biochemistry and Molecular Biology.
Figure 4. Editing of homocysteine, aminoacylation of thiols and synthesis of ‐Hcy‐tRNA by MetRS. (a) The MetRS‐catalysed cyclisation of homocysteinyl adenylate to form Hcy‐thiolactone and AMP, which are subsequently released from the synthetic/editing active site of MetRS. (b) The MetRS‐catalysed reaction of a thiol (mimicking the side chain of Hcy, R‐CH2SH) with methionyl‐tRNA to form a methionine thioester, which is subsequently released from the synthetic/editing active site of MetRS. (c) MetRS‐catalysed aminoacylation of tRNA with ‐Hcy. Reproduced with permission from Jakubowski (2011) © Acta Biochimica Polonica.
Figure 6. The synthetic/editing active site of MetRS: Hydrophobic and hydrogen bonding interactions provide specificity for the cognate substrate l‐methionine. Superimposition of carbon chain backbones for the MetRS·Met complex (burly‐wood) and free MetRS (light grey), solved at 1.8 Å resolution, shows movements of active site residues upon binding of methionine. Residue colours are red in the MetRS·Met complex and green in free MetRS, and l‐methionine is magenta. Reprinted from Serre et al. (2001). © Elsevier.
Figure 7. (a) Conformations of pretransfer substrate analogue, NvaAMS, bound in the synthetic and editing sites of LeuRS. (b) Locations of NvaAMS in the synthetic and editing active sites of LeuRS. Reprinted from Lincecum et al. (2003) © Elsevier.
Figure 8. Metabolism of Hcy in human endothelial cells. Hcy arises from the dietary protein methionine as a by‐product of cellular transmethylation reactions. Hcy is metabolised by MetRS to Hcy‐thiolactone, which is enhanced when re‐methylation to methionine, catalysed by methionine synthase (MS), is impaired, for example, by inadequate folate or vitamin 12 supply. Hcy‐thiolactone diffuses out and into the cell, reacts with protein lysine residues or is hydrolysed to Hcy by an HDL‐associated serum Hcy‐thiolactonase/PON1. Hcy forms a mixed disulphide with serum albumin (Hcy‐ ‐protein). A reaction with nitric oxide, produced by endothelial nitric oxide synthase, converts Hcy to ‐Hcy ( ‐NO‐Hcy), which is then incorporated translationally into protein following formation of ‐Hcy‐tRNA catalysed by MetRS. Hcy‐ ‐protein contains Hcy in amide or peptide bonds. Reprinted from Jakubowski (2002). © The American Society for Biochemistry and Molecular Biology.
Figure 9. Editing and aminoacylation sites of ThrRS. (a) Crystal structure of ThrRS editing site (PDB: 1TJE). C182 is close to H73, H77 and H186. All four residues are critical for post‐transfer editing of Ser‐tRNAThr. (b) Crystal structure of ThrRS aminoacylation site (PDB: 1EVK). C334, H385 and H511 coordinate a zinc ion, which is essential for tRNAThr aminoacylation. (c) Sequence alignment of conserved motifs in ThrRS editing and aminoacylation sites. : ; : ; : ; : ; : . (d) A model for ThrRS editing site C182 oxidation. Two nitrogen atoms of H73 and H186 stabilise the deprotonated C182 thiolate nucleophile that attacks the oxygen atom of an oxidant. X = OH, Cl. Reproduced with permission from Wu et al. (2014) © Oxford University Press.
Figure 10. LeuRS, a sensor of amino acids, transmits a signal to TORC1. (a) In yeast and mammalian cells, leucine‐loaded LeuRS, which is required for protein synthesis, binds and activates Gtr/Rag GTPases, which in turn activate TORC1. Activated TORC1 then facilitates protein synthesis while turning off the protein recycling process, autophagy. (b) A comparison between TORC1 activation in yeast and in mammalian cells. In yeast, LeuRS binds and maintains GTP‐bound Gtr1, while in mammalian cells, LeuRS binds RagD and acts as a GAP that maintains the GDP‐bound RagD. Reproduced with permission form Segev and Hay (2012) © Elsevier.
Figure 2. Alternative editing pathways for non‐cognate amino acids. A cognate amino acid proceeds through the aminoacylation pathway, indicated by the double‐headed arrows, to form AA‐tRNA. A non‐cognate amino acid enters the aminoacylation pathway but is rejected at points indicated by single‐headed arrows (Jakubowski and Fersht, ).
Figure 5. ‐Homocysteinylation of protein lysine residues by Hcy‐thiolactone (Jakubowski, , ).


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Further Reading

Chwatko G, Boers GH, Strauss KA, Shih DM and Jakubowski H (2007) Mutations in methylenetetrahydrofolate reductase or cystathionine beta‐synthase gene, or a high‐methionine diet, increase homocysteine thiolactone levels in humans and mice. Federation of American Societies for Experimental Biology Journal 21: 1707–1713.

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Jakubowski H (2012) Quality control in tRNA charging. Wiley Interdisciplinary Reviews: RNA 3: 295–310.

Jakubowski H and Goldman E (1992) Editing of errors in selection of amino acids for protein synthesis. Microbiological Reviews 56: 412–429.

Ling J, Reynolds N and Ibba M (2009) Aminoacyl‐tRNA synthesis and translational quality control. Annual Review of Microbiology 63: 61–78.

O'Donoghue P, Ling J, Wang YS and Söll D (2013) Upgrading protein synthesis for synthetic biology. Nature Chemical Biology 9: 594–598.

Pang YL, Poruri K and Martinis SA (2014) tRNA synthetase: tRNA aminoacylation and beyond. Wiley Interdisciplinary Reviews: RNA 5: 461–480.

Perona JJ and Gruic‐Sovulj I (2014) Synthetic and editing mechanisms of aminoacyl‐tRNA synthetases. Topics in Current Chemistry 344: 1–42.

Yadavalli SS and Ibba M (2012) Quality control in aminoacyl‐tRNA synthesis its role in translational fidelity. Advances in Protein Chemistry and Structural Biology 86: 1–43.

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Jakubowski, Hieronim(Mar 2015) Transfer RNA Synthetase Editing of Errors in Amino Acid Selection. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0000532.pub3]