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Literature summary for 6.3.2.2 extracted from
- Identification of an important cysteine residue in human glutamate-cysteine ligase catalytic subunit by site-directed mutagenesis (1998), Biochem. J., 336, 675-680.
No PubMed abstract available
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expression of the His-tagged wild-type enzyme and mutants in Spodoptera frugiperda Sf9 cells via baculovirus infection | Homo sapiens |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| C248G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
| C249G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, reduced activity of the holoenzyme compared to the wild-type enzyme | Homo sapiens |
| C295G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
| C491G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
| C501G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
| C52G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
| C553G | site-directed mutagenesis in the catalytic subunit, slightly reduced activity of the catalytic subunit, about 3.5fold reduced activity of the holoenzyme compared to the wild-type enzyme | Homo sapiens |
| C605G | site-directed mutagenesis in the catalytic subunit, reduced activity of the catalytic subunit, activity of the holoenzyme is similar to the wild-type enzyme | Homo sapiens |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| cysteamine | inactivation of wild-type enzyme and mutant C553G after 90 min at 4°C, 0.2 mM cysteamine and 2 mM ATP | Homo sapiens |  |
| dithiothreitol | inhibition of wild-type holoenzyme and C553G mutant holoenzyme, the latter is more sensitive | Homo sapiens | |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.5 | - |
L-cysteine | recombinant mutant C553G holoenzyme, pH 8.0, 37°C | Homo sapiens | |
| 0.7 | - |
L-Glu | recombinant wild-type holoenzyme, pH 8.0, 37°C | Homo sapiens | |
| 0.8 | - |
L-cysteine | recombinant wild-type holoenzyme, pH 8.0, 37°C | Homo sapiens | |
| 0.9 | - |
L-Glu | recombinant mutant C553G holoenzyme, pH 8.0, 37°C | Homo sapiens | |
| 3.6 | - |
L-2-aminobutyrate | recombinant wild-type holoenzyme, pH 8.0, 37°C | Homo sapiens | |
| 5 | - |
L-2-aminobutyrate | recombinant mutant C553G holoenzyme, pH 8.0, 37°C | Homo sapiens | |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 75000 | - |
2 * 75000, recombinant wild-type and mutant enzymes, SDS-PAGE | Homo sapiens |
| 114000 | - |
recombinant wild-type and mutant holoenzymes, gel filtration | Homo sapiens |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ATP + L-Glu + L-Cys | Homo sapiens | rate-limiting step in glutathione biosynthesis | ADP + phosphate + gamma-L-Glu-L-Cys | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Homo sapiens | - |
- |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His-tagged wild-type and mutant enzymes from Sf9 insect cells | Homo sapiens |
Reaction
| Reaction | Comment | Organism | Reaction ID |
|---|---|---|---|
| ATP + L-glutamate + L-cysteine = ADP + phosphate + gamma-L-glutamyl-L-cysteine | Cys553 is not the active site residue | Homo sapiens |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 0.12 | - |
purified recombinant mutant C249G catalytic subunit and mutant C295G catalytic subunit | Homo sapiens |
| 0.17 | - |
purified recombinant mutant C52G catalytic subunit and mutant C248G catalytic subunit | Homo sapiens |
| 0.38 | - |
purified recombinant mutant C605G catalytic subunit | Homo sapiens |
| 0.48 | - |
purified recombinant mutant C501G catalytic subunit | Homo sapiens |
| 0.63 | - |
purified recombinant mutant C491G catalytic subunit | Homo sapiens |
| 0.92 | - |
purified recombinant mutant C553G catalytic subunit | Homo sapiens |
| 1.15 | - |
purified recombinant wild-type catalytic subunit | Homo sapiens |
| 1.83 | - |
purified recombinant mutant C553G holoenzyme | Homo sapiens |
| 4.67 | - |
purified recombinant mutant C249G holoenzyme | Homo sapiens |
| 5.17 | - |
purified recombinant mutant C605G holoenzyme | Homo sapiens |
| 5.33 | - |
purified recombinant mutant C295G holoenzyme | Homo sapiens |
| 5.67 | - |
purified recombinant mutant C248G holoenzyme | Homo sapiens |
| 6.17 | - |
purified recombinant wild-type holoenzyme | Homo sapiens |
| 6.67 | - |
purified recombinant mutant C52G holoenzyme | Homo sapiens |
| 7 | - |
purified recombinant mutants C491G holoenzyme and C501G holoenzyme | Homo sapiens |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ATP + L-Glu + L-2-aminobutyrate | - |
Homo sapiens | ADP + phosphate + L-Glu-2-aminobutyrate | - |
? | |
| ATP + L-Glu + L-Cys | reaction can be performed by the catalytic subunit alone, but presence of the regulatory subunit in the holoenzyme increases the activity | Homo sapiens | ADP + phosphate + gamma-L-Glu-L-Cys | - |
? | |
| ATP + L-Glu + L-Cys | rate-limiting step in glutathione biosynthesis | Homo sapiens | ADP + phosphate + gamma-L-Glu-L-Cys | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| dimer | 2 * 75000, recombinant wild-type and mutant enzymes, SDS-PAGE | Homo sapiens |
| More | reaction can be performed by the catalytic subunit alone, but presence of the regulatory subunit in the holoenzyme increases the activity | Homo sapiens |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| GLCL | - |
Homo sapiens |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 37 | - |
assay at | Homo sapiens |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 8 | - |
assay at | Homo sapiens |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| ATP | - |
Homo sapiens | |
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