Glutathione and its Related Enzymes in the Nile Fish |
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| Authors: | Ragaa R Hamed Tahany M Maharem Rasha A M Guinidi |
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| Institution: | (1) Department of Molecular Biology, National Research Centre, Cairo, Egypt;(2) Department of Biochemistry, Ain-Shams University, Egypt |
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| Abstract: | Glutathione (GSH) and related enzymes, glutathione transferase (GST), glutathione peroxidase (GPx) and glutathione reductase
(GR) form an important phase 2 biotransformation enzymes system. The objective of this study was to compare this enzymes system
in three fish species from the river Nile, Oreochromis niloticus, Claris lazera and Cyprinus carpio in order to establish the main differences and to purify and characterize GST from the liver of O. niloticus.The level of GSH and the activity of GST, GPx and GR in the liver, kidney and gills of the three fish species were examined.
A simple reproducible procedure for the purification of GST from the liver of O. niloticus to homogeneity, which includes chromatography on DEAE- cellulose followed by affinity chromatography on GSH-sepharose was
established. The molecular mass was found to be 25,460 Da by SDS-PAGE. The Michaelis-Meneten constants (Km) of the enzyme for GSH and CDNB were 0.35 mM and 0.42 mM, respectively. The affinity purified enzyme exhibited maximum pH
at pH 8.0 and increasing pH above 8.0 did not affect the observed maximum. The purified enzyme acts readily on CDNB, less
readily on some standard transferase substrates (1,2-dichloro-4-nitrobenzene and p-nitrophenethyl bromide) and not at all on others (bromosulphophthalein and p-nitrobenzyl chloride). Bromosulfophthalein, cibacron blue and hematin inhibited CDNB-conjugating activity of the purified
enzyme with IC50 0.079, 3.98 and 0.126 μM, respectively. |
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| Keywords: | glutathione peroxidase glutathione reductase glutathione-S-transferase and purification |
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