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Literature summary for 6.1.1.1 extracted from

  • Raina, M.; Moghal, A.; Kano, A.; Jerums, M.; Schnier, P.D.; Luo, S.; Deshpande, R.; Bondarenko, P.V.; Lin, H.; Ibba, M.
    Reduced amino acid specificity of mammalian tyrosyl-tRNA synthetase is associated with elevated mistranslation of Tyr codons (2014), J. Biol. Chem., 289, 17780-17790 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 6.1.1.1

Cloned(Commentary)

Cloned (Comment) Organism
gene EGW00102, sequence comparisons, recombinant expression of codon-optimized His6-tagged enzyme in Escherichia coli strain BL21(DE3), recombinant expression of the enzyme ligated to a ribozyme for in vitro transcription by T7 polymerase Cricetulus griseus

Protein Variants

Protein Variants Comment Organism
A74G site-directed mutagenesis, replacement of Ala74 with Gly increases the Km 7fold and has no effect on kcat for Tyr. The kcat/Km for Phe decreases 5fold Cricetulus griseus
D122N site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
G120N site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
H77T site-directed mutagenesis, mutation of His77 to the smaller nonhydrophobic Thr increases the Km and decreases the kcat by 40fold Cricetulus griseus
L125W site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
additional information generation of a truncated mutant enzyme TyrRS Cricetulus griseus
N82D site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
W40C site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
Y123W site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus
Y52H site-directed mutagenesis, mutant substrate specificity compared to the wild-type enzyme Cricetulus griseus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information Michaelis-Menten steady-state kinetics. Steady-state kinetic constants for ATP-[32P]PPi exchange for CHO cytosolic full length wild-type and variant TyrRS Cricetulus griseus
0.146
-
 L-tyrosine recombinant wild-type enzyme, pH 7.8, 25°C Cricetulus griseus
0.16
-
 L-tyrosine recombinant truncated mutant enzyme, pH 7.8, 25°C Cricetulus griseus

Localization

Localization Comment Organism GeneOntology No. Textmining
cytosol
-
 Cricetulus griseus 5829
-

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Cricetulus griseus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + L-tyrosine + tRNATyr Cricetulus griseus
-
 AMP + diphosphate + L-tyrosyl-tRNATyr
-
 ?

Organism

Organism UniProt Comment Textmining
Cricetulus griseus G3H935
-
-

Source Tissue

Source Tissue Comment Organism Textmining
CHO cell
-
 Cricetulus griseus
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + L-tyrosine + tRNATyr
-
 Cricetulus griseus AMP + diphosphate + L-tyrosyl-tRNATyr
-
 ?
ATP + L-tyrosine + tRNATyrGTA two sequences, tRNATyr1 and tRNATyr2, substrate CHO tRNATyrGTA is recombinantly expressed. CHOTyrRS is able to attach Phe to both tRNATyr2 and CHO total tRNA containing native tRNATyr, excluding the possibility that mischarging results from the lack of post-transcriptional modifications to the in vitro-transcribed substrate Cricetulus griseus AMP + diphosphate + L-tyrosyl-tRNATyrGTA
-
 ?
additional information the presence of the C-terminal EMAP II-domain has no effect on the recognition of cognate Tyr and discrimination against noncognate Phe Cricetulus griseus ?
-
 ?

Synonyms

Synonyms Comment Organism
CHOTyrRS
-
 Cricetulus griseus
Tyrosyl-tRNA synthetase
-
 Cricetulus griseus
TyrRS
-
 Cricetulus griseus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25 37 aminoacylation assay at Cricetulus griseus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
12.5
-
 L-tyrosine recombinant wild-type enzyme, pH 7.8, 25°C Cricetulus griseus
15
-
 L-tyrosine recombinant truncated mutant enzyme, pH 7.8, 25°C Cricetulus griseus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.8
-
 aminoacylation assay at Cricetulus griseus

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Cricetulus griseus

General Information

General Information Comment Organism
evolution mammalian TyrRS has evolved to be significantly less accurate than its bacterial counterpart. Different evolutionary constraints determine the accuracy of translation quality control in eukaryotes and bacteria. Functional comparisons of mammalian and bacterial tyrosyl-tRNA synthetase reveal key differences at residues responsible for amino acid recognition, highlighting differences in evolutionary constraints for translation quality control, overview Cricetulus griseus
malfunction reduced amino acid specificity of mammalian tyrosyl-tRNA synthetase is associated with elevated mistranslation of Tyr codons. Mischarging of tRNATyr with noncognate Phe by tyrosyl-tRNA synthetase is responsible for mistranslation. Steady-state kinetic analyses of CHO cytoplasmic tyrosyl-tRNA synthetase reveals a 25fold lower specificity for Tyr over Phe as compared with previously characterized bacterial enzymes, consistent with the observed increase in translation error rates during tyrosine limitation Cricetulus griseus
additional information residues, not conserved between bacteria and eukaryotes, Cys35, His48, Thr51, and Lys233 in bacterial TyrRS interact with ATP during transition state formation. Comparison the Geobacillus stearothermophilus, PDB ID 1tyd, and human, PDB ID 1q11, TyrRS active sites, overview. Hydrogen-bonding interactions between bacterial TyrRS residues Asp176 and Tyr34 and the substrate Tyr hydroxyl group help confer amino acid specificity to the enzyme Cricetulus griseus
physiological function aa-tRNA synthesis is a two-step reaction: activation of an amino acid with ATP to form aminoacyl adenylate, followed by transfer of the aminoacyl moiety to the 3' end of the tRNA. The error rate of this first step of translation is largely dependent on the specificity of the aaRS, that is selection of the correct amino acid and tRNA from the respective cellular pools of predominantly noncognate substrates. aaRSs select their cognate tRNAs by exploiting sequence-specific differences between various tRNAs during binding and aminoacylation. Translation of Tyr codons is highly prone to Phe isincorporation during amino acid limitation inCHOcells. CHO cell TyrRS is error-prone and readily aminoacylates tRNATyr with Phe, cf. EC 6.1.1.20. Steady-state kinetic analyses of CHO cytoplasmic tyrosyl-tRNA synthetase reveals a 25fold lower specificity for Tyr over Phe as compared with previously characterized bacterial enzymes, consistent with the observed increase in translation error rates during tyrosine limitation Cricetulus griseus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
93.75
-
 L-tyrosine recombinant truncated mutant enzyme, pH 7.8, 25°C Cricetulus griseus
856.16
-
 L-tyrosine recombinant wild-type enzyme, pH 7.8, 25°C Cricetulus griseus