Literature summary for 4.1.1.15 extracted from

Fan, L.Q.; Li, M.W.; Qiu, Y.J.; Chen, Q.M.; Jiang, S.J.; Shang, Y.J.; Zhao, L.M.
Increasing thermal stability of glutamate decarboxylase from Escherichia coli by site-directed saturation mutagenesis and its application in GABA production (2018), J. Biotechnol., 278, 1-9 .

Search for more literature for 4.1.1.15

Cloned(Commentary)

Cloned (Comment)	Organism
gene gadB, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain DH5alpha	Escherichia coli

Protein Variants

Protein Variants	Comment	Organism
additional information	application of site-directed saturation mutagenesis of the N-terminal residues of GadB from Escherichia coli to improve its thermostability. Among the mutants tested, M6 is the most thermostable one	Escherichia coli
Q5I/V6D/T7Q	mutant M6, site-directed mutagenesis, the mutant shows higher thermostability, with a 5.6times (560%) increase in half-life value at 45°C, 8.7°C rise in melting temperature (Tm) and a 14.3°C rise in the temperature at which 50% of the initial activity remained after 15 min incubation (T15/50), compared to wild-type enzyme. The induced new hydrogen bonds in the same polypeptide chain or between polypeptide chains in Escherichia coli GadB homohexamer may be responsible for the improved thermostability. Increased thermostability contributes to increased GABA conversion ability. After 12 h conversion of 3 mol/l glutamate, 297 g/l GABA is produced and 95% mole conversion rate is catalyzed by mtant M6 whole cells while those by wild-type GAD are 273.5 g/L and 86.2%, respectively	Escherichia coli
Q5N/V6Y/T7V	mutant M1, site-directed mutagenesis, the mutant shows improved thermostability and increased activity compared to the wild-type enzyme	Escherichia coli
Q5Y/V6R/T7K	mutant M8, site-directed mutagenesis, the mutant shows improved thermostability and increased activity compared to the wild-type enzyme	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	A5YKJ2	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and desalting gel filtration, to over 90% purity	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	three-dimensional enzyme structure analysis	Escherichia coli

Synonyms

Synonyms	Comment	Organism
GadB	-	Escherichia coli

Temperature Optimum [°C]

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

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
40	-	after 12 h incubation at 40°C, the residual activity of mutants M1, M6 and M8 is 52.05%, 65.08% and 30.2%, respectively, about 4.16, 5.2 and 2.4fold that of wild-type GadB (12.5%)	Escherichia coli
45	-	half-life of recombinant wild-type GadB is 24.24 min, while the half-lives of mutants M1, M6 and M8 are 128.84 min, 160.45 min and 126.03 min, respectively. The half-lives of M1, M6 and M8 are 5.32, 6.62 and 5.20fold that of the wild-type enzyme	Escherichia coli
50	60	after 12 h incubation at 50°C and 60°C, the residual activity of wild-type GadB is 12.5% and 9.0%, respectively. The residual activity of mutants M1, M6 and M8 is 35.16% and 27.17%, 38.10% and 33.25%, 27.19% and 24.39%, respectively, which is at least 2fold higher than that of wild-type GadB	Escherichia coli
100	-	boiling for 5 min inactivates the enzyme	Escherichia coli

pH Optimum

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

Cofactor

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

General Information

General Information	Comment	Organism
additional information	three-dimensional enzyme structure analysis, homology modeling using the crystal structure of homohexameric GadB at low pH, PDB ID 1pmm	Escherichia coli

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Release 2023.1 (January 2023)