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The action of sodium metavanadate with erythrocytes as studied by 1H spin-echo and 51V NMR spectroscopy.
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Abstracts
1011 THE ACTION OF SODIUM METAVANADATE WITH ERYTHROCYTES AS STUDIED BY lH SPIN-ECHO AND 51V NMR SPECTROSCOPY. M. Garner, J. Reglinski and W.E. Smith Department of Pure and Applied Chemistry, University of Strathclyde,
Glasgow, Gl lXL, U.K. Vanadate is known to be an inhibitor of Na/K ATPase by binding to the aspartate 369 residue, which is the site of phosphorylation. Vanadate may also be regarded as a mild oxidant and has been shown to be reduced by glutathione in erythrocytes [ 11. Spin echo NMR spectroscopy is a technique which can be used to assess glutathione directly in the cell [2]. The method of assessment compares resonances within the spectrum. One resonance in particular is derived from the hydrogen environment adjacent to the thiol function of glutathione; this resonance is therefore susceptible to changes in the oxidation state of glutathione. Therefore, using this method, it is possible to monitor both the total glutathione and the oxidised/reduced glutathione redox balance of the cell. TTn:+., c,allI& ‘II spin echo and 51x,’WMR spectroscopy the action of sodium metavanadate on intact human erythrocytes was probed. Millimolar concentrations of vanadate were shown to cause a gradual depletion of total intracellular glutathione and a slight depletion of intracellular ergothionine. However, the oxidised/reduced glutathione redox balance remained constant over this period. After blocking the erythrocyte anion transporter with 4,4’-diisothiocyanatostilbene-2,2’-disulfonic acid (DIDS) and treating with vanadate, the total glutathione level and redox balance remain relatively constant. These results suggest that DIDS blocks the transport of vanadate into the cell via the anion transporter and that glutathione is capable of modifying vanadate prior to coupling at the Na/K ATPase binding site without altering the glutathione redox balance. References 1) LG. Macara, K. Kustin and L.C. Cantley, Jr, Biochim. Biophys. Acta, 629, 9.5 (1980) 2) D.L. Rabenstein, J. Biochem. Biophys. Methods, 9, 277 (1984)
Abstracts
1011 THE ACTION OF SODIUM METAVANADATE WITH ERYTHROCYTES AS STUDIED BY lH SPIN-ECHO AND 51V NMR SPECTROSCOPY. M. Garner, J. Reglinski and W.E. Smith Department of Pure and Applied Chemistry, University of Strathclyde,
Glasgow, Gl lXL, U.K. Vanadate is known to be an inhibitor of Na/K ATPase by binding to the aspartate 369 residue, which is the site of phosphorylation. Vanadate may also be regarded as a mild oxidant and has been shown to be reduced by glutathione in erythrocytes [ 11. Spin echo NMR spectroscopy is a technique which can be used to assess glutathione directly in the cell [2]. The method of assessment compares resonances within the spectrum. One resonance in particular is derived from the hydrogen environment adjacent to the thiol function of glutathione; this resonance is therefore susceptible to changes in the oxidation state of glutathione. Therefore, using this method, it is possible to monitor both the total glutathione and the oxidised/reduced glutathione redox balance of the cell. TTn:+., c,allI& ‘II spin echo and 51x,’WMR spectroscopy the action of sodium metavanadate on intact human erythrocytes was probed. Millimolar concentrations of vanadate were shown to cause a gradual depletion of total intracellular glutathione and a slight depletion of intracellular ergothionine. However, the oxidised/reduced glutathione redox balance remained constant over this period. After blocking the erythrocyte anion transporter with 4,4’-diisothiocyanatostilbene-2,2’-disulfonic acid (DIDS) and treating with vanadate, the total glutathione level and redox balance remain relatively constant. These results suggest that DIDS blocks the transport of vanadate into the cell via the anion transporter and that glutathione is capable of modifying vanadate prior to coupling at the Na/K ATPase binding site without altering the glutathione redox balance. References 1) LG. Macara, K. Kustin and L.C. Cantley, Jr, Biochim. Biophys. Acta, 629, 9.5 (1980) 2) D.L. Rabenstein, J. Biochem. Biophys. Methods, 9, 277 (1984)