Literature summary for 2.3.2.2 extracted from

Terzyan, S.; Cook, P.; Heroux, A.; Hanigan, M.
Structure of 6-diazo-5-oxo-norleucine-bound human gamma-glutamyl transpeptidase 1, a novel mechanism of inactivation (2017), Protein Sci., 26, 1196-1205 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene GGT1, recombinant expression of enzyme mutant V272A in Pichia pastoris strain X-33	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant deglycosylated hGGT1 mutant V272A with bound inhibitor DON, mixing of 0.002 ml of 4.3 mg/ml protein in 50 mM HEPES, pH 8.0, 0.5 mM EDTA, 0.02% sodium azide, and 2 mM DON, with 0.0017 ml of H2O, and 0.002 ml of reservoir solution containing 20-25% PEG 3350, 0.1 M Na-cacodylate, pH 6.0, and 0.1 M ammonium chloride, microseeding with apo-hGGT1 crystals, room temperature, 1-2 days, X-ray diffraction structure determination and analysis at 2.20 A resolution, modeling using the apo-form crystals of hGGT1 (PDB ID 4Z9O) as template	Homo sapiens

Crystallization (Comment)

Organism

purified recombinant deglycosylated hGGT1 mutant V272A with bound inhibitor DON, mixing of 0.002 ml of 4.3 mg/ml protein in 50 mM HEPES, pH 8.0, 0.5 mM EDTA, 0.02% sodium azide, and 2 mM DON, with 0.0017 ml of H2O, and 0.002 ml of reservoir solution containing 20-25% PEG 3350, 0.1 M Na-cacodylate, pH 6.0, and 0.1 M ammonium chloride, microseeding with apo-hGGT1 crystals, room temperature, 1-2 days, X-ray diffraction structure determination and analysis at 2.20 A resolution, modeling using the apo-form crystals of hGGT1 (PDB ID 4Z9O) as template

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
V272A	structure analysis	Homo sapiens

Inhibitors

Inhibitors	Comment	Organism
6-diazo-5-oxo-L-norleucine	i.e. DON, a glutamate analogue, irreversible inhibitor of hGGT1, but also inhibits many essential glutamine metabolizing enzymes rendering it too toxic for use in the clinic as a GGT1 inhibitor. Binding structure analysis with human gamma-glutamyl transpeptidase 1, and molecular mechanism of hGGT1 inactivation by DON, overview. The crystal structure of DON-inactivated hGGT1 contains a molecule of DON without the diazo-nitrogen atoms in the active site. The overall structure of the hGGT1-DON complex resembles the structure of the apo-enzyme. The structure of hGGT1-DON complex reveals two covalent bonds between the enzyme and inhibitor which are part of a six membered ring. The ring includes the OG atom of Thr381, the reactive nucleophile of hGGT1 and the alpha-amine of Thr381. The mechanism of inactivation by DON differs from its inactivation of other glutamine metabolizing enzymes	Homo sapiens
GGsTop	a glutamate analog that inactivates hGGT1 by forming one covalent and 11 hydrogen bonds with the enzyme, increasing the melting temperature by 20°C	Homo sapiens
additional information	identification of inhibitors of the enzyme GGT1	Homo sapiens

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
1.2	-	L-gamma-glutamyl-4-nitroanilide	pH 7.4, 37°C, recombinant enzyme	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
extracellular	-	Homo sapiens	-	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
a (5-L-glutamyl)-peptide + an amino acid	Homo sapiens	-	a peptide + a 5-L-glutamyl amino acid	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	P19440	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant GGT1 mutant V272A from Pichia pastoris strain X-33	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
a (5-L-glutamyl)-peptide + an amino acid	-	Homo sapiens	a peptide + a 5-L-glutamyl amino acid	-	?
L-gamma-glutamyl-4-nitroanilide + glycylglycine	-	Homo sapiens	4-nitroaniline + 5-L-glutamyl-glycylglycine	-	?

Synonyms

Synonyms	Comment	Organism
gamma-glutamyl transferase 1	-	Homo sapiens
gamma-glutamyl transpeptidase 1	-	Homo sapiens
GGT1	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Homo sapiens

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
additional information	-	melting temperature of enzyme GGT1 is 53-58°C depending on the buffer system and pH used, while with bound inhibitor 6-diazo-5-oxo-L-norleucine, the mellting temperature is 69-72°C, overview. The unbound enzyme is most stable in Na-citrate, pH 6.7, the inhibitor-bound enzyme in HEPES, pH 7.0	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.4	-	assay at	Homo sapiens

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
2.7	-	6-diazo-5-oxo-L-norleucine	pH 7.4, 37°C, recombinant enzyme	Homo sapiens

General Information

General Information	Comment	Organism
additional information	hGGT1 catalytic reaction mechanism, activation of the catalytic nucleophile that initiates the hGGT1 reaction, overview	Homo sapiens
physiological function	enzyme gamma-glutamyl transpeptidase 1 (GGT1) cleaves extracellular gamma-glutamyl compounds and contributes to the pathology of asthma, reperfusion injury and cancer	Homo sapiens