Literature summary for 6.1.1.20 extracted from

Sasaki, H.M.; Sekine, S.; Sengoku, T.; Fukunaga, R.; Hattori, M.; Utsunomiya, Y.; Kuroishi, C.; Kuramitsu, S.; Shirouzu, M.; Yokoyama, S.
Structural and mutational studies of the amino acid-editing domain from archaeal/eukaryal phenylalanyl-tRNA synthetase (2006), Proc. Natl. Acad. Sci. USA, 103, 14744-14749.

Cloned(Commentary)

Cloned (Comment)	Organism
expression of N-terminal fragment of the PheRS beta-subunit in Escherichia coli strain BL21(DE3), expression of selenomethionine-labeled N-terminal fragment of the PheRS beta-subunit in Escherichia coli strain B834(DE3)	Pyrococcus horikoshii

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant selenomethionine-labeled N-terminal fragment of the PheRS beta-subunit, 0.001 ml of protein solution containing 36 mg/ml protein in 20 mM Tris-HCl buffer, pH 8.0, containing 200 mM NaCl, mixed with 0.001 ml of precipitant solution, containing 100 mM sodium citrate, pH 5.6, and 16.5% PEG 20000, covarage with a 1:1 mixture of paraffin oil and silicone, 20°C, 1 week, soaking in a soaked in a cryoprotectant solution containing 100 mM sodium citrate, pH 5.6, 18.2% PEG 20000, and 20% glycerol, X-ray diffraction structure determination and analysis at 1.94 A resolution	Pyrococcus horikoshii

Crystallization (Comment)

Organism

purified recombinant selenomethionine-labeled N-terminal fragment of the PheRS beta-subunit, 0.001 ml of protein solution containing 36 mg/ml protein in 20 mM Tris-HCl buffer, pH 8.0, containing 200 mM NaCl, mixed with 0.001 ml of precipitant solution, containing 100 mM sodium citrate, pH 5.6, and 16.5% PEG 20000, covarage with a 1:1 mixture of paraffin oil and silicone, 20°C, 1 week, soaking in a soaked in a cryoprotectant solution containing 100 mM sodium citrate, pH 5.6, 18.2% PEG 20000, and 20% glycerol, X-ray diffraction structure determination and analysis at 1.94 A resolution

Pyrococcus horikoshii

Protein Variants

Protein Variants	Comment	Organism
A141W	site-directed mutagenesis, the mutant exhibits high tyrosine mischarging activity	Pyrococcus horikoshii
D234A	site-directed mutagenesis, the mutant exhibits moderate tyrosine mischarging activity, the mutant PheRS incorrectly hydrolyze the cognate Phe-tRNAPhe	Pyrococcus horikoshii
E127A	site-directed mutagenesis, the mutant exhibits low tyrosine mischarging activity	Pyrococcus horikoshii
E219A	site-directed mutagenesis, the mutant is similar to the wild-type enzyme	Pyrococcus horikoshii
F145A	site-directed mutagenesis, the mutant is similar to the wild-type enzyme	Pyrococcus horikoshii
I216A	site-directed mutagenesis, the mutant is similar to the wild-type enzyme	Pyrococcus horikoshii
L168A	site-directed mutagenesis, the mutant exhibits moderate tyrosine mischarging activity and shows reduced Tyr-tRNAPhe deacylation activity	Pyrococcus horikoshii
L202A	site-directed mutagenesis, the mutant PheRS incorrectly hydrolyze the cognate Phe-tRNAPhe	Pyrococcus horikoshii
L210A	site-directed mutagenesis, the mutant is similar to the wild-type enzyme	Pyrococcus horikoshii
N217A	site-directed mutagenesis, the mutant exhibits high tyrosine mischarging activity and shows abolished Tyr-tRNAPhe deacylation activity	Pyrococcus horikoshii
Q126A	site-directed mutagenesis, the mutant shows reduced Tyr-tRNAPhe deacylation activity	Pyrococcus horikoshii
R137A	site-directed mutagenesis, the mutant exhibits low tyrosine mischarging activity and shows reduced Tyr-tRNAPhe deacylation activity	Pyrococcus horikoshii
R223A	site-directed mutagenesis, the mutant exhibits moderate tyrosine mischarging activity and shows reduced Tyr-tRNAPhe deacylation activity	Pyrococcus horikoshii
S211A	site-directed mutagenesis, the mutant PheRS incorrectly hydrolyze the cognate Phe-tRNAPhe	Pyrococcus horikoshii
T221A	site-directed mutagenesis, the mutant is similar to the wild-type enzyme	Pyrococcus horikoshii
T236A	site-directed mutagenesis, the mutant PheRS incorrectly hydrolyze the cognate Phe-tRNAPhe	Pyrococcus horikoshii
Y189A	site-directed mutagenesis, the mutant exhibits low tyrosine mischarging activity	Pyrococcus horikoshii

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	-	Pyrococcus horikoshii

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
ATP + L-phenylalanine + tRNAPhe	Pyrococcus horikoshii	-	AMP + diphosphate + L-phenylalanyl-tRNAPhe	-	?

Organism

Organism	UniProt	Comment	Textmining
Pyrococcus horikoshii	O73984	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N-terminal fragment of the PheRS beta-subunit from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, recombinant selenomethionine-labeled N-terminal fragment of the PheRS beta-subunit from Escherichia coli strain B834(DE3) by anion exchange chromatography, adsorption chromatography, and gel filtration	Pyrococcus horikoshii

Reaction

Reaction	Comment	Organism	Reaction ID
ATP + L-phenylalanine + tRNAPhe = AMP + diphosphate + L-phenylalanyl-tRNAPhe	residue N217 is essential for catalysis	Pyrococcus horikoshii	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
ATP + L-phenylalanine + tRNAPhe	-	Pyrococcus horikoshii	AMP + diphosphate + L-phenylalanyl-tRNAPhe	-	?
ATP + L-phenylalanine + tRNAPhe	editing mechanism of noncognate aminoacyl-tRNA involving domains B3 and B4 and residues Leu202, Ser211, Asp234, and Thr236, overview	Pyrococcus horikoshii	AMP + diphosphate + L-phenylalanyl-tRNAPhe	-	?

Subunits

Subunits	Comment	Organism
More	the N-terminal fragment of the PheRS beta-subunit includes the editing domain B3/4, which has archaea/eukarya-specific insertions/deletions and adopts a different orientation relative to other domains, as compared with that of bacterial PheRS, structure, overview	Pyrococcus horikoshii

Synonyms

Synonyms	Comment	Organism
Phenylalanyl-tRNA synthetase	-	Pyrococcus horikoshii
PheRS	-	Pyrococcus horikoshii

Cofactor

Cofactor	Comment	Organism	Structure
ATP	-	Pyrococcus horikoshii