Literature summary for 2.3.2.2 extracted from

Hibi, T.; Imaoka, M.; Shimizu, Y.; Itoh, T.; Wakayama, M.
Crystal structure analysis and enzymatic characterization of gamma-glutamyltranspeptidase from Pseudomonas nitroreducens (2019), Biosci. Biotechnol. Biochem., 83, 262-269 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene ggt, recombinant overexpression of wild-type and mutant enzymes	Pseudomonas nitroreducens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type PnGGT enzyme, sitting drop vapor diffusion method, mixing of 0.001 ml of 3-5 mg/ml protein in 0.1 M HEPES, pH 7.0, with 0.001 ml reservoir solution containing 16% PEG 8000, 15% PEG 400, 0.1 M HEPES, pH 7.0, 50 mM glycylglycine, and equilibration against 0.5 ml of reservoir solution, 20°C, 1 week, method optimization, X-ray diffraction structure determination and analysis at 1.57-1.70 A resolution, molecular replacement method using the structure of Escherichia coli GGT (EcGGT, PDB ID 2DG5) as the template, modeling	Pseudomonas nitroreducens

Crystallization (Comment)

Organism

purified recombinant wild-type PnGGT enzyme, sitting drop vapor diffusion method, mixing of 0.001 ml of 3-5 mg/ml protein in 0.1 M HEPES, pH 7.0, with 0.001 ml reservoir solution containing 16% PEG 8000, 15% PEG 400, 0.1 M HEPES, pH 7.0, 50 mM glycylglycine, and equilibration against 0.5 ml of reservoir solution, 20°C, 1 week, method optimization, X-ray diffraction structure determination and analysis at 1.57-1.70 A resolution, molecular replacement method using the structure of Escherichia coli GGT (EcGGT, PDB ID 2DG5) as the template, modeling

Pseudomonas nitroreducens

Protein Variants

Protein Variants	Comment	Organism
F417Y	site-directed mutagenesis, the mutant shows decreased hydrolysis and increased transfer activity, i.e. gamma-glutamyl-p-nitroanilide hydrolysis and gamma-L-glutamylhydroxamate synthesis, compared to wild-type	Pseudomonas nitroreducens
W385T	site-directed mutagenesis, the mutant shows decreased hydrolysis and increased transfer activity, i.e. gamma-glutamyl-p-nitroanilide hydrolysis and gamma-L-glutamylhydroxamate synthesis, compared to wild-type	Pseudomonas nitroreducens
W525A	site-directed mutagenesis, the mutant shows decreased hydrolysis and increased transfer activity, i.e. gamma-glutamyl-p-nitroanilide hydrolysis and gamma-L-glutamylhydroxamate synthesis, compared to wild-type	Pseudomonas nitroreducens

Inhibitors

Inhibitors	Comment	Organism	Structure
2-amino-4-([3-(carboxymethyl)phenoxy](methoyl)phosphoryl)butanoic acid	peptidyl phosphonate inhibitor GGsTop, the hydroxy-2-oxoethylamino-oxidanylidene-butane moiety of the inhibitor mimicks an acceptor substrate	Pseudomonas nitroreducens

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas nitroreducens	E0D5C2	-	-
Pseudomonas nitroreducens IFO12694	E0D5C2	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type and mutant enzymes	Pseudomonas nitroreducens

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
7.1	-	about, recombinant wild-type enzyme, pH 10.5, 30°C, transpeptidation activity	Pseudomonas nitroreducens
15	-	about, recombinant enzyme mutant W385T, pH 10.5, 30°C, transpeptidation activity	Pseudomonas nitroreducens
16	-	about, recombinant enzyme mutant W525A, pH 10.5, 30°C, transpeptidation activity	Pseudomonas nitroreducens
17	-	about, recombinant enzyme mutant F417Y, pH 10.5, 30°C, transpeptidation activity	Pseudomonas nitroreducens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
gamma-glutamyl-4-nitroanilide + hydroxamate	-	Pseudomonas nitroreducens	4-nitroaniline + gamma-L-glutamylhydroxamate	-	?
gamma-glutamyl-4-nitroanilide + hydroxamate	-	Pseudomonas nitroreducens IFO12694	4-nitroaniline + gamma-L-glutamylhydroxamate	-	?
additional information	the gamma-glutamyltranspeptidase (GGT) from Pseudomonas nitroreducens strain IFO12694 (PnGGT) has a unique preference for primary amines as gamma-glutamyl acceptors over standard L-amino acids and peptides, detection of the structural basis of PnGGT's hydrolysis and transpeptidation reactions, overview. Key interactions between three residues Trp385, Phe417, and Trp525 distinguish PnGGT from other GGTs, the aromatic side chains of Trp385, Phe417, and Trp525 are involved in the recognition of acceptor substrates. Local structures of substrate binding sites	Pseudomonas nitroreducens	?	-	-
additional information	the gamma-glutamyltranspeptidase (GGT) from Pseudomonas nitroreducens strain IFO12694 (PnGGT) has a unique preference for primary amines as gamma-glutamyl acceptors over standard L-amino acids and peptides, detection of the structural basis of PnGGT's hydrolysis and transpeptidation reactions, overview. Key interactions between three residues Trp385, Phe417, and Trp525 distinguish PnGGT from other GGTs, the aromatic side chains of Trp385, Phe417, and Trp525 are involved in the recognition of acceptor substrates. Local structures of substrate binding sites	Pseudomonas nitroreducens IFO12694	?	-	-

Synonyms

Synonyms	Comment	Organism
gamma-glutamyltranspeptidase	-	Pseudomonas nitroreducens
GGT	-	Pseudomonas nitroreducens
More	see also EC 3.4.19.13	Pseudomonas nitroreducens
PnGGT	-	Pseudomonas nitroreducens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Pseudomonas nitroreducens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
10.5	-	assay at	Pseudomonas nitroreducens

General Information

General Information	Comment	Organism
additional information	comparisons of the active site structures of GGT enzyme from Pseudomonas nitroreducens with those from Escherichia coli and Homo sapiens, overview. The residues around the donor substrate-binding site of PnGGT (Arg94, Asn384, Glu403, Asp406, Ser435, Ser436, Gly456, and Gly456) are conserved among the other GGTs. In the active site of PnGGT, Phe417 in the 411-421 loop and Trp385 form the side wall of the postulated acceptor-binding pocket	Pseudomonas nitroreducens