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Literature summary for 4.1.1.15 extracted from

  • Jun, C.; Joo, J.C.; Lee, J.H.; Kim, Y.H.
    Thermostabilization of glutamate decarboxylase B from Escherichia coli by structure-guided design of its pH-responsive N-terminal interdomain (2014), J. Biotechnol., 174, 22-28.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
gene gadB, expression of C-terminally His-tagged wild-type and mutant enzyme in Escherichia coli strain BL21(DE3), cloning in Escherichia coli strain DH5alpha Escherichia coli

Protein Variants

Protein Variants Comment Organism
additional information improvement of the thermostability of GadB through structural optimization ofits N-terminal interdomain. Residues Gln5, Val6, and Thr7 are potential mutational target sites for the optimization of inter- and intra-molecular interactions of the triple-helix bundle. Generation of a deletion mutant of GadB lacking residues 1-14 at the N-terminus, GadBDELTA1-14 Escherichia coli
Q5D/V6I/T7E site-directed mutagenesis the mutant shows higher thermostability and increased melting temperature compared to the wild-type, but shows no reduction of catalytic activity Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.32
-
L-glutamate pH 4.6, 37°C, recombinant untagged wild-type enzyme Escherichia coli
20.31
-
L-glutamate pH 4.6, 37°C, recombinant C-terminally His-tagged wild-type enzyme Escherichia coli
25.09
-
L-glutamate pH 4.6, 37°C, recombinant deletion mutant GadBDELTA1-14 Escherichia coli
26.02
-
L-glutamate pH 4.6, 37°C, recombinant mutant Q5D/V6I/T7E Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Cl- binding of chloride ions to the N-terminus could stabilizes the triple-helix bundle Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-glutamate Escherichia coli
-
4-aminobutanoate + CO2
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli P69910 gene gadB
-

Purification (Commentary)

Purification (Comment) Organism
recombinant C-terminally His-tagged wild-type and mutant enzyme from Escherichia coli strain BL21(DE3) by affinity chromatography and ultrafiltration Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-glutamate
-
Escherichia coli 4-aminobutanoate + CO2
-
?

Subunits

Subunits Comment Organism
homohexamer
-
Escherichia coli
More homohexameric GadB forms a triple-helix bundle interdomain at acidic pH Escherichia coli

Synonyms

Synonyms Comment Organism
GadB
-
Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Escherichia coli

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
additional information
-
homohexameric GadB forms a triple-helix bundle interdomain at acidic pH and contributes to the thermostability of GadB Escherichia coli
55
-
purified recombinant wild-type enzyme, without chloride ions, 10 min, 71% activity remaining at pH 4.6, 2% remaining at pH 8.0, the deletion mutant GadB DELTA1-14 exhibits similar thermostabilityat pH 4.6 and pH 8.0 Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
59.26
-
L-glutamate pH 4.6, 37°C, recombinant C-terminally His-tagged wild-type enzyme Escherichia coli
73.35
-
L-glutamate pH 4.6, 37°C, recombinant deletion mutant GadBDELTA1-14 Escherichia coli
75.41
-
L-glutamate pH 4.6, 37°C, recombinant mutant Q5D/V6I/T7E Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
4.6
-
assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
pyridoxal 5'-phosphate
-
Escherichia coli

General Information

General Information Comment Organism
additional information the N-terminal fourteen residues (1-14) of homohexameric GadB forms a triple-helix bundle interdomain at acidic pH and contributes to the thermostability of GadB as the pH shifts from pH 7.6 to pH 4.6 Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.88
-
L-glutamate pH 4.6, 37°C, recombinant mutant Q5D/V6I/T7E Escherichia coli
2.92
-
L-glutamate pH 4.6, 37°C, recombinant C-terminally His-tagged wild-type enzyme and deletion mutant GadBDELTA1-14 Escherichia coli