Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Brekken, D.L.; Phillips, M.A.
    Trypanosoma brucei gamma-glutamylcysteine synthetase. Characterization of the kinetic mechanism and the role of Cys-319 in cystamine inactivation (1998), J. Biol. Chem., 273, 26317-26322.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
6.3.2.2 expression of the C-terminally His8-tagged wild-type enzyme in Escherichia coli BL21(DE3), expression of the mutant C319A in Escherichia coli BO265 Trypanosoma brucei
6.3.2.2 expression of wild-type and mutant C319A enzymes in Escherichia coli BL21(DE3) as His-tagged proteins Trypanosoma brucei

Protein Variants

EC Number Protein Variants Comment Organism
6.3.2.2 C319A mutant is insensitive to cystamine, but the catalytic efficiency, the kinetic mechanism, or the substrate affinities remain unaltered Trypanosoma brucei
6.3.2.2 C319A site-directed mutagenesis, loss of sensitivity against cysteamine inactivation Trypanosoma brucei

Inhibitors

EC Number Inhibitors Comment Organism Structure
6.3.2.2 cystamine irreversible inactivation of the wild-type enzyme, loss of 75% activity within 10 min at 0.01 mM, binds to active site Cys319 and in this way blocks the binding of substrate to the enzyme, no inhibition of the mutant C319A enzyme Trypanosoma brucei
6.3.2.2 cysteamine wild-type enzyme is inhibited by 75% at 0.01 mM after 10 min, complete irreversible inhibition at 10 mM, the mutant C319Ais completely resistant to cysteamine Trypanosoma brucei

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.3.2.2 additional information
-
 additional information kinetics Trypanosoma brucei
6.3.2.2 additional information
-
 additional information kinetics, wild-type enzyme and mutant C319A Trypanosoma brucei
6.3.2.2 1.4
-
 ATP wild-type enzyme and mutant C319A, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 2.6
-
 L-Glu wild-type enzyme, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 5.4
-
 L-2-aminobutyric acid wild-type enzyme, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 5.9
-
 L-Glu mutant C319A, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 6.1
-
 L-2-aminobutyric acid mutant C319A, pH 8.0, 37°C Trypanosoma brucei

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.3.2.2 Mg2+
-
 Trypanosoma brucei

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6.3.2.2 ATP + L-Glu + L-Cys Trypanosoma brucei first step in the biosynthesis of trypanothione via GSH ADP + phosphate + gamma-L-Glu-L-Cys
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
6.3.2.2 Trypanosoma brucei
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
6.3.2.2 recombinant His-tagged wild-type and mutant C319A enzymes from Escherichia coli Trypanosoma brucei
6.3.2.2 recombinant mutant C319A and recombinant His8-tagged wild-type from Escherichia coli Trypanosoma brucei

Reaction

EC Number Reaction Comment Organism Reaction ID
6.3.2.2 ATP + L-glutamate + L-cysteine = ADP + phosphate + gamma-L-glutamyl-L-cysteine rapid equilibrium random ter-reactant mechanism, substrate binding modeling, Cys319 is an active site cysteine not essential for activity Trypanosoma brucei
a
6.3.2.2 ATP + L-glutamate + L-cysteine = ADP + phosphate + gamma-L-glutamyl-L-cysteine reaction and kinetic mechanism, modeling Trypanosoma brucei
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6.3.2.2 ATP + L-Glu + L-2-aminobutyrate
-
 Trypanosoma brucei ADP + phosphate + gamma-L-Glu-2-aminobutyrate
-
 ?
6.3.2.2 ATP + L-Glu + L-alpha-aminobutyrate
-
 Trypanosoma brucei ADP + phosphate + gamma-L-Glu-L-alpha-aminobutyrate
-
 ?
6.3.2.2 ATP + L-Glu + L-Cys
-
 Trypanosoma brucei ADP + phosphate + gamma-L-Glu-L-Cys
-
 ?
6.3.2.2 ATP + L-Glu + L-Cys first step in the biosynthesis of trypanothione via GSH Trypanosoma brucei ADP + phosphate + gamma-L-Glu-L-Cys
-
 ?
6.3.2.2 additional information binding of ATP to the enzyme increases the binding affinity for L-Glu by 18fold, while binding of L-Glu or L-alpha-aminobutyrate decreases the affinity for binding of the other by 6fold Trypanosoma brucei ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
6.3.2.2 gamma-GCS
-
 Trypanosoma brucei
6.3.2.2 gamma-Glutamylcysteine synthetase
-
 Trypanosoma brucei

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
6.3.2.2 37
-
 assay at Trypanosoma brucei

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.3.2.2 23
-
 ATP wild-type and mutant enzyme, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 23
-
 L-Glu wild-type and mutant enzyme, pH 8.0, 37°C Trypanosoma brucei
6.3.2.2 23
-
 L-alpha-aminobutyrate wild-type and mutant enzyme, pH 8.0, 37°C Trypanosoma brucei

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
6.3.2.2 8
-
 assay at Trypanosoma brucei

Cofactor

EC Number Cofactor Comment Organism Structure
6.3.2.2 ATP
-
 Trypanosoma brucei
6.3.2.2 ATP binding of L-Glu to the enzyme increases the binding affinity for ATP by 18fold Trypanosoma brucei