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Literature summary for 6.1.1.1 extracted from

  • Kiga, D.; Sakamoto, K.; Kodama, K.; Kigawa, T.; Matsuda, T.; Yabuki, T.; Shirouzu, M.; Harada, Y.; Nakayama, H.; Takio, K.; Hasegawa, Y.; Endo, Y.; Hirao, I.; Yokoyama, S.
    An engineered Escherichia coli tyrosyl-tRNA synthetase for site-specific incorporation of an unnatural amino acid into proteins in eukaryotic translation and its application in a wheat germ cell-free system (2002), Proc. Natl. Acad. Sci. USA, 99, 9715-9720.
    View publication on PubMedView publication on EuropePMC

Application

Application Comment Organism
biotechnology site-specific incorporation of 3-iodo-L-tyrosine into proteins in a cell-free system for use in specialized in vitro translation systems Escherichia coli

Cloned(Commentary)

Cloned (Comment) Organism
overexpression of His-tagged wild-type and mutants in strain BL21(DE3) Escherichia coli

Protein Variants

Protein Variants Comment Organism
Q179A site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q179E inactive mutant Escherichia coli
Q179N site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q179S site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q179Y inactive mutant Escherichia coli
Q195A site-directed mutagenesis, active with L-tyrosine, low activity with 3-iodo-L-tyrosine Escherichia coli
Q195C site-directed mutagenesis, reduced activity with L-tyrosine, low activity with 3-iodo-L-tyrosine Escherichia coli
Q195D site-directed mutagenesis, highly reduced activity with L-tyrosine, low activity with 3-iodo-L-tyrosine Escherichia coli
Q195E site-directed mutagenesis, active with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195F inactive mutant Escherichia coli
Q195G site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195H site-directed mutagenesis, active with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195I site-directed mutagenesis, highly reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195K inactive mutant Escherichia coli
Q195L site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195M site-directed mutagenesis, highly reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195N site-directed mutagenesis, reduced activity with L-tyrosine, low activity with 3-iodo-L-tyrosine Escherichia coli
Q195R inactive mutant Escherichia coli
Q195S site-directed mutagenesis, active with L-tyrosine, low activity with 3-iodo-L-tyrosine Escherichia coli
Q195T site-directed mutagenesis, active with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195V site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Q195W inactive mutant Escherichia coli
Q195Y site-directed mutagenesis, reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Y37A/Q195A inactive mutant Escherichia coli
Y37A/Q195C site-directed mutagenesis, active with L-tyrosine and 3-iodo-L-tyrosine, preference for the latter, reduced overall activity Escherichia coli
Y37A/Q195N inactive mutant Escherichia coli
Y37A/Q195S site-directed mutagenesis, equally low activity with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y37I/Q195A site-directed mutagenesis, equally low activity with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y37I/Q195C inactive mutant Escherichia coli
Y37I/Q195N inactive mutant Escherichia coli
Y37I/Q195S inactive mutant Escherichia coli
Y37L/Q195A site-directed mutagenesis, highly reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Y37L/Q195C site-directed mutagenesis, equally low activity with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y37L/Q195N site-directed mutagenesis, highly reduced activity with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Y37L/Q195S inactive mutant Escherichia coli
Y37V/Q195N site-directed mutagenesis, active with L-tyrosine and 3-iodo-L-tyrosine, preference for the latter, reduced overall activity Escherichia coli
Y37V/Q195S site-directed mutagenesis, reduced activity with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y73A site-directed mutagenesis, equally active with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y73F site-directed mutagenesis, active with L-tyrosine, no activity with 3-iodo-L-tyrosine Escherichia coli
Y73G site-directed mutagenesis, equally active with L-tyrosine and 3-iodo-L-tyrosine, reduced overall activity Escherichia coli
Y73H site-directed mutagenesis, active with L-tyrosine, and slightly active with 3-iodo-L-tyrosine Escherichia coli
Y73I site-directed mutagenesis, active with L-tyrosine and 3-iodo-L-tyrosine, preference for the latter Escherichia coli
Y73L site-directed mutagenesis, equally active with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y73M site-directed mutagenesis, equally active with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y73S site-directed mutagenesis, active with L-tyrosine and 3-iodo-L-tyrosine, preference for the first Escherichia coli
Y73V site-directed mutagenesis, equally active with L-tyrosine and 3-iodo-L-tyrosine Escherichia coli
Y73V/Q195A site-directed mutagenesis, active with L-tyrosine and 3-iodo-L-tyrosine, preference for the latter, reduced overall activity Escherichia coli
Y73V/Q195C site-directed mutagenesis, 10fold more active with 3-iodo-L-tyrosine than with L-tyrosine, reduced overall activity Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0053
-
L-tyrosine pH 7.6, 37°C, wild-type enzyme Escherichia coli
0.13
-
3-iodo-L-tyrosine pH 7.6, 37°C, mutant Y73V/Q195C Escherichia coli
0.14
-
L-tyrosine pH 7.6, 37°C, mutant Y73V/Q195C Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + L-tyrosine + tRNATyr Escherichia coli
-
AMP + L-Tyr-tRNATyr + diphosphate
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from strain BL21(DE3) Escherichia coli

Reaction

Reaction Comment Organism Reaction ID
ATP + L-tyrosine + tRNATyr = AMP + diphosphate + L-tyrosyl-tRNATyr amino acid residues Y37 and Q195 are involved in substrate specificity determination Escherichia coli

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
-
Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + 3-iodo-L-tyrosine + tRNATyr mutant Y73V/Q195C and other mutants, no activity with the wild-type enzyme Escherichia coli AMP + L-Tyr-tRNATyr + diphosphate
-
?
ATP + L-tyrosine + tRNATyr
-
Escherichia coli AMP + L-Tyr-tRNATyr + diphosphate
-
?

Synonyms

Synonyms Comment Organism
Tyrosyl-tRNA synthetase
-
Escherichia coli
TyrRS
-
Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.045
-
L-tyrosine pH 7.6, 37°C, mutant Y73V/Q195C Escherichia coli
0.43
-
3-iodo-L-tyrosine pH 7.6, 37°C, mutant Y73V/Q195C Escherichia coli
12
-
L-tyrosine pH 7.6, 37°C, wild-type enzyme Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.6
-
assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
ATP
-
Escherichia coli