Cloned (Comment) | Organism |
---|---|
overexpression of His-tagged wild-type and mutant D-aminoacyl-tRNA deacylase-like domains in Escherichia coli strain BL21(DE3), mutant M129K is located in inclusion bodies | Pyrococcus abyssi |
Crystallization (Comment) | Organism |
---|---|
purified recombinant D-aminoacyl-tRNA deacylase-like domain with bound Ser3AA, hanging drop vapor diffusion method, mixing of equal volumes of protein, ligand and reservoir solution, crystallization from 0.2 M (NH4)2SO4, 0.1 M sodium cacodylate, pH 6.5, and 30% PEG 8000 and 0.1 M HEPES, pH 7.0, and 25% PEG 3350, purified recombinant D-aminoacyl-tRNA deacylase-like domain with bound SerAMS, using 0.1 M Bis-Tris, pH 6.5, and 25% PEG 3350, and the serine Pab-NTDL-serine cocrystals from 0.1 M HEPES pH 7.0 and 25% PEG 8000, X-ray diffraction structure determination and analysis at 1.86-2.25 A resolution, overview | Pyrococcus abyssi |
Protein Variants | Comment | Organism |
---|---|---|
E134A | site-directed mutagenesis, the mutation has no effect on the deacylation activity | Pyrococcus abyssi |
H83A | site-directed mutagenesis, the mutant possesses the editing activity albeit with a slower rate compared to the wild-type enzyme | Pyrococcus abyssi |
K121A | no expression for the alanine mutant | Pyrococcus abyssi |
K121M | site-directed mutagenesis, substitution of Lys121 to serine does not abolish Ser-tRNAThr deacylation activity | Pyrococcus abyssi |
K121S | site-directed mutagenesis, substitution of Lys121 to serine results in a complete abolition of Ser-tRNAThr deacylation activity | Pyrococcus abyssi |
M129K | site-directed mutagenesis, the mutant shows binding not only to L-serine but also to a variety of other L-amino acids that are tested in addition to binding to various D-amino acids, overview | Pyrococcus abyssi |
Y120A | site-directed mutagenesis, the mutation has no effect on the deacylation activity | Pyrococcus abyssi |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | - |
Pyrococcus abyssi |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + L-threonine + tRNAThr | Pyrococcus abyssi | - |
AMP + diphosphate + L-threonyl-tRNAThr | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pyrococcus abyssi | Q9UZ14 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and mutant D-aminoacyl-tRNA deacylase-like domains from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration | Pyrococcus abyssi |
Renatured (Comment) | Organism |
---|---|
recombinant mutant M129K D-aminoacyl-tRNA deacylase-like domain from inclusion bodies after expression in Escherichia coli strain BL21(DE3) in 0.1 M phosphate buffer at pH 8.0 containing 6 M guanidinium hydrochloride and 10 mM Tris-HCl, followed by nickel affinity chromatography, the denatured protein is refolded stepwise by dialysis in 0.1 M phosphate buffer, pH 8.0, containing 10 mM Tris-HCl | Pyrococcus abyssi |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + L-threonine + tRNAThr | - |
Pyrococcus abyssi | AMP + diphosphate + L-threonyl-tRNAThr | - |
? | |
ATP + L-threonine + tRNAThr | the main chains atoms of Tyr119 and Tyr120 are sufficient to prevent the deacylation of Thr-tRNAThr | Pyrococcus abyssi | AMP + diphosphate + L-threonyl-tRNAThr | - |
? | |
additional information | mischarging of the enzyme with noncognate amino acids, overview, post-transfer editing mechanism of the D-aminoacyl-tRNA deacylase-like domain in the archaeal threonyl-tRNA synthetase, mechanistic insights into the removal of noncognate L-serine from tRNAThr, M129 is responsible for enantiomeric selection in DTD, Glu134 is involved in fixing the seryl moiety in the active site, overview | Pyrococcus abyssi | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
Threonyl-tRNA synthetase | - |
Pyrococcus abyssi |
ThrRS | - |
Pyrococcus abyssi |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Pyrococcus abyssi |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.2 | - |
asay at | Pyrococcus abyssi |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
ATP | - |
Pyrococcus abyssi |