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Purification and characterization of γ-glutamyl transpeptidase from Ascaris suum
γ-Glutamyl transpeptidase, which is of central importance in the degradation of glutathione, was purified from Ascaris suum to apparent homogeneity. The enzyme was found to be a predominantly membrane-bound protein and was solubilized by Triton X-100. The purified enzyme, which exhibits a specific a...
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Published in: | Molecular and biochemical parasitology 1996-04, Vol.77 (1), p.41-47 |
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Main Authors: | , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that cite this one |
Online Access: | Get full text |
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Summary: | γ-Glutamyl transpeptidase, which is of central importance in the degradation of glutathione, was purified from
Ascaris suum to apparent homogeneity. The enzyme was found to be a predominantly membrane-bound protein and was solubilized by Triton X-100. The purified enzyme, which exhibits a specific activity of 1009 U (mg protein)
−1, showed a molecular mass of 70 kDa and was found to be composed of two non-identical subunits of molecular mass 43 and 30 kDa. Concerning the kinetic properties of the enzyme, the data presented in this study showed that various amino acids and dipeptides with L-configuration served as acceptors for the γ-glutamyl moieties of the enzyme reaction products and showed
K
m-values in the mM range. The apparent
K
m-value for the γ-glutamyl donor L-glutamyl-γ-7-amido-4-methylcoumarin of the enzyme was found to be 0.03 mM. L- and D-serine in combination with borate ions were competitive inhibitors of the enzyme activity with
K
i-values of 0.30 and 0.61 mM, respectively. Acivicin was an irreversible inhibitor of the enzyme with a
K
i-value of 0.42 mM and with a pseudo-first-order kinetics (
k
inact) of 0.18 min
−1. In vitro treatment of the adult
A. suum with acivicin resulted in a dose-dependent inhibition of the enzyme activity and an increase of the glutathione levels. These findings indicate the physiological role of the γ-glutamyl transpeptidase of this parasitic nematode in the catabolism of glutathione. |
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ISSN: | 0166-6851 1872-9428 |
DOI: | 10.1016/0166-6851(96)02573-X |