Cloned (Comment) | Organism |
---|---|
overexpression of His-tagged AsnRS in Escherichia coli strain DH5alpha, the enzyme is toxic when heterologously overexpressed in Escherichia coli, because of sequestration of tRNAAsn as Asp-tRNAAsn, this toxicity is rescued upon coexpression of the Helicobacter pylori Asp/Glu-Adt, EC 6.3.5.6 | Helicobacter pylori |
Protein Variants | Comment | Organism |
---|---|---|
L81N | site-directed mutagenesis, the mutation in the anticodon binding domain doubles the kcat for tRNAAsn as compared to the wild-type enzyme | Helicobacter pylori |
L81N/L86M | site-directed mutagenesis, the mutation in the anticodon binding domain alters the tRNA specificity as compared to the wild-type enzyme, the L81N/L86M mutant does not follow Michaelis-Menten kinetics | Helicobacter pylori |
L86M | site-directed mutagenesis, the mutation in the anticodon binding domain alters the tRNA specificity as compared to the wild-type enzyme | Helicobacter pylori |
additional information | mutations in the anticodon binding domain of Helicobacter pylori ND-AspRS, e.g. at L81, L86, N82, and M87, reduce this enzymes ability to misacylate tRNAAsn and enhance tRNAAsp specificity, in a manner that correlates with the toxicity of the enzyme in Escherichia coli, overview | Helicobacter pylori |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | the L81N/L86M mutant does not follow Michaelis-Menten kinetics | Helicobacter pylori | |
0.77 | - |
tRNAAsp | pH 7.5, recombinant wild-type enzyme | Helicobacter pylori | |
0.83 | - |
tRNAAsn | pH 7.5, recombinant wild-type enzyme | Helicobacter pylori | |
0.85 | - |
tRNAAsp | pH 7.5, recombinant mutant L81N enzyme | Helicobacter pylori | |
0.87 | - |
tRNAAsp | pH 7.5, recombinant mutant L86M enzyme | Helicobacter pylori | |
1.87 | - |
tRNAAsn | pH 7.5, recombinant mutant L81N enzyme | Helicobacter pylori | |
2.23 | - |
tRNAAsn | pH 7.5, recombinant mutant L86M enzyme | Helicobacter pylori |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | - |
Helicobacter pylori |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + L-aspartate + tRNAAsn | Helicobacter pylori | - |
AMP + diphosphate + aspartyl-tRNAAsn | - |
? | |
ATP + L-aspartate + tRNAAsp | Helicobacter pylori | - |
AMP + diphosphate + aspartyl-tRNAAsp | - |
? | |
additional information | Helicobacter pylori | in bacteria that lack AsnRS, AspRS is nondiscriminating and generates both Asp-tRNAAsp and the noncanonical, misacylated Asp-tRNAAsn, this misacylated tRNA is subsequently repaired by the glutamine-dependent Asp-tRNAAsn/Glu-tRNAGln amidotransferase, EC 6.3.5.6, increasing tRNAAsp specificity in an ND-AspRS diminishes in vivo toxicity | ? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Helicobacter pylori | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged AsnRS from Escherichia coli strain DH5alpha by nickel affinity chromatography | Helicobacter pylori |
Storage Stability | Organism |
---|---|
-20°C, purified recombinant enzyme, in 50% glycerol, and 33 mM phosphate, pH 7.4, 3 mM Tris-HCl, 1.5 mM 2-mercaptoethanol, and 0.5 mM phenylmethanesulfonyl fluoride, stable for months | Helicobacter pylori |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + L-aspartate + tRNAAsn | - |
Helicobacter pylori | AMP + diphosphate + aspartyl-tRNAAsn | - |
? | |
ATP + L-aspartate + tRNAAsn | recombinantly produced tRNA substrate | Helicobacter pylori | AMP + diphosphate + aspartyl-tRNAAsn | - |
? | |
ATP + L-aspartate + tRNAAsp | - |
Helicobacter pylori | AMP + diphosphate + aspartyl-tRNAAsp | - |
? | |
ATP + L-aspartate + tRNAAsp | recombinantly produced tRNA substrate | Helicobacter pylori | AMP + diphosphate + aspartyl-tRNAAsp | - |
? | |
additional information | in bacteria that lack AsnRS, AspRS is nondiscriminating and generates both Asp-tRNAAsp and the noncanonical, misacylated Asp-tRNAAsn, this misacylated tRNA is subsequently repaired by the glutamine-dependent Asp-tRNAAsn/Glu-tRNAGln amidotransferase, EC 6.3.5.6, increasing tRNAAsp specificity in an ND-AspRS diminishes in vivo toxicity | Helicobacter pylori | ? | - |
? | |
additional information | tRNA anticodon binding site structures, overview, Helicobacter pylori AspRS is a nondiscriminating enzyme that aminoacylates both tRNAAsp and tRNAAsn, ND-AspRS is 1.7times more efficient at aminoacylating tRNAAsp over tRNAAsn | Helicobacter pylori | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
ND-AspRS | - |
Helicobacter pylori |
nondiscriminating aspartyl-tRNA synthetase | - |
Helicobacter pylori |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | the wild-type enzyme has a kcat for tRNAAsp that is 60% higher than that of tRNAAsn | Helicobacter pylori | |
0.014 | - |
tRNAAsn | pH 7.5, recombinant wild-type enzyme | Helicobacter pylori | |
0.015 | - |
tRNAAsn | pH 7.5, recombinant mutant L86M enzyme | Helicobacter pylori | |
0.021 | - |
tRNAAsp | pH 7.5, recombinant mutant L86M enzyme | Helicobacter pylori | |
0.022 | - |
tRNAAsp | pH 7.5, recombinant wild-type enzyme | Helicobacter pylori | |
0.026 | - |
tRNAAsn | pH 7.5, recombinant mutant L81N enzyme | Helicobacter pylori | |
0.028 | - |
tRNAAsp | pH 7.5, recombinant mutant L81N enzyme | Helicobacter pylori |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Helicobacter pylori |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
ATP | - |
Helicobacter pylori |