Direct Effect of CD on Glutathione S-Transferase and Glutathione Reductase from Calystegia Sepium

Abstract

Interactions between heavy metals, glutathione, glutathione S-transferase (GST), and glutathione reductase (GR) are being investigated by many working groups, but evaluation of the direct effect of Cd²⁺ on these enzymes in vitro is lacking. We report here the effect of cadmium (10, 50, 100, 250 μM CdSO₄) on partially purified enzymes from Calystegia sepium. Plants were grown under normal field conditions without metals and the enzymes were extracted by Tris buffer and partially purified by ammonium sulphate fractionation and gel filtration. Glutathione S-transferase activity was measured with different substrates, i.e., 1–chloro-2,4-dinitrobenzene (CDNB), p-nitrobenzylchloride (NBC), and the herbicide Fluorodifen. GST activity was significantly lower in leaf compared to stem, flower, and rhizome and the inhibitory effect of Cd was obtained with NBC and Fluorodifen substrates at 250 μM. There was no effect of Cd on GR activity up to 250 μM.

KEY WORDS:

• Calystegia sepium
• heavy metals
• glutathione
• glutathione S-transferase
• CdSO_4, cadmium

ACKNOWLEDGEMENTS

The authors thank Frank Laturnus, Linköpings Universitet Sweden, for critically reading the manuscript. Funding for Lyudmila Lyubenova by a grant from the Bavarian State Ministry for Education and Arts in the frame of the BAYHOST program is gratefully accepted.