Incorporation of labelled glycine into reduced glutathione of intact human erythrocytes by enzyme-catalysed exchange. A nuclear-magnetic-resonance study

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Abstract
1H, 2H and 15N NMR spectroscopy was used to monitor the incorporation of free glycine into the glycine residue of reduced glutathione (GSH) in suspensions of intact human erythrocytes. By using 1H spin-echo NMR the exchange reaction between [2H5]glycine and the protonated glycine residue of GSH was studied at various [2H5]glycine concentrations, thus enabling the calculation of an apparent Km and Vmax for the process. The reaction is catalyzed by glutathione synthetase and proceeds most rapidly in the absence of glucose, which is the main physiological energy source of the erythrocyte. 15N NMR spectroscopy, with a 1-pulse sequence, and 2H NMR spectroscopy, with an inversion recovery method, enabled demonstration of the incorporation of labeled glycine into an intraerythrocyte peptide, consistent with incorporation into GSH. The exchange reaction, although inhibited by glucose, appeared not to be dependent on low ATP or 2,3-bisphosphoglycerate concentrations.