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3.4.19.13: glutathione gamma-glutamate hydrolase

This is an abbreviated version!
For detailed information about glutathione gamma-glutamate hydrolase, go to the full flat file.

Word Map on EC 3.4.19.13

Reaction

a glutathione-S-conjugate
+
H2O
=
an (L-cysteinylglycine)-S-conjugate
 +
L-glutamate

Synonyms

At4g29210, BaGGT42, BaGGT469, BlGGT13, BsGGT168, gamma-glutamyl transpeptidase, gamma-glutamyl-transpeptidase, gamma-GT, GGT, GGT-1, GGT1, GGT3, GGT4, GGT5, hp1118, More

ECTree

     3 Hydrolases
         3.4 Acting on peptide bonds (peptidases)
             3.4.19 Omega peptidases
                3.4.19.13 glutathione gamma-glutamate hydrolase

Crystallization

Crystallization on EC 3.4.19.13 - glutathione gamma-glutamate hydrolase

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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
hanging-drop vapor diffusion method, X-ray diffraction data for the co-crystal of transpeptidase and acivicin at 1.45 A resolution. SUMO-tag renders high expression and solubility
in complex with acivicin, to 1.8 A resolution. Acivicin is bound covalently through its C3 atom with sp2 hybridization to Thr403 Ogamma, the catalytic nucleophile of the enzyme
mutant T391A, unable to undergo autocatalytic processing, at 2.55 A resolution. Structural comparison of the precursor and mature protein demonstrates that the structures of the core regions in the two proteins are unchanged, with marked differences near the active site. In the precursor, the segment corresponding to the C-terminal region of the L-subunit occupies the site where the loop, residues 438-449, forms the lid of the gamma-glutamyl group-binding pocket in the mature enzyme. Upon cleavage of the N-terminal peptide bond of Thr391, the newly produced C-terminus, residues 375–390, flips out, allowing the 438-449 segment to form the gamma-glutamyl group-binding pocket. A water molecule is located near the carbonyl carbon atom of Gln-390. The spatial arrangement around the water and Thr391 relative to the scissile peptide bond appears suitable for the initiation of autocatalytic processing
to 1.95 A resolution. The enzyme has a stacked alphabetabetaalpha fold comprising the large and small subunits, similar to the folds seen in members of the N-terminal nucleophile hydrolase superfamily. The active site residue Thr391, the N-terminal residue of the small subunit, is located in the groove, from which the pocket for gamma-glutamyl moiety binding follows. The gamma-glutamyl-enzyme intermediate and the structure of the complex with L-glutamate reveal how the gamma-glutamyl moiety and L-glutamate are recognized by the enzyme. A water molecule is seen on the carbonyl carbon of the gamma-glutamyl-Thr391 O bond in the intermediate that is to be hydrolyzed. The residues essential for enzymic activity, i.e. Arg114, Asp433, Ser462, and Ser463, are all involved in the binding of the gamma-glutamyl moiety
X-ray crystallography of the enzyme in complex with inhibitor 2-amino-4-[[3-(carboxymethyl)phenoxy](methoyl)phosphoryl] butanoic acid
to 1.9 A resolution. The refined model contains two 40-kDa/20-kDa heterodimers in the asymmetric unit and has structural features comparable with other N-terminal nucleophile hydrolases. Autoprocessing of the enzyme leads to a large conformational change, with the loop preceding the catalytic residue Thr380 moving more than 35 A, thus relieving steric constraints that likely limit substrate binding. Cleavage of the proenzyme results in the formation of a threonine-threonine dyad comprised of Thr380 and Thr398. The hydroxyl group of Thr398 is located equidistant from the alpha-amino group and hydroxyl side chain of Thr380
crystals of are grown at room temperature by vapor diffusion with the hanging drop method. For crystallization studies, the natural variant V272A of the human enzyme is expressed in Pichia pastoris strain X-33, purified, and deglycosylated. Crystal structures of human enzyme, including the free enzyme, inhibitor-bound transition states, and glutamate-bound enzyme. Crystal structures of human gamma-glutamyl transpeptidase show conformational changes within the active site as the enzyme progresses from the free enzyme to inhibitor bound tetrahedral transition states and finally to the glutamate bound structure prior to the release of this final product of the reaction. The structure of the apoenzyme shows flexibility within the active site. The serine-borate-bound enzyme crystal structure demonstrates that serine-borate occupies the active site of the enzyme, resulting in an enzyme-inhibitor complex that replicates the enzyme's tetrahedral intermediate/transition state. The structure of 2-amino-4-[[3-(carboxymethyl)phenyl](methyl)phosphono]-butanoic acid-bound enzyme reveals its interactions with the enzyme and why neutral phosphonate diesters are more potent inhibitors than monoanionic phosphonates