R. Stephen Reid

A proton nuclear magnetic resonance study of the binding of methylmercury in human erythrocytes

The binding of methylmercury, CH3Hg(II), by small molecules in the intracellular region of human erythrocytes has been studied by 1H-NMR spectroscopy. To suppress or completely eliminate interfering resonances from the much more abundant hemoglobin protons, spectra were measured by a technique based on the transfer of saturation throughout the envelope of hemoglobin resonances following a selective presaturation pulse or by the spin-echo Fourier transform method. With these techniques, 1H-NMR spectra were measured for the more abundant intracellular small molecules, including glycine, alanine, creatine, lactic acid, ergothioneine and glutathione, both intact and hemolyzed erythrocytes to which CH3Hg(II) had been added. The results for intact erythrocytes indicate that part of the CH3Hg(II) is complexed by intracellular glutathione. These results also indicate that exchange of CH3Hg(II) among glutathione molecules is fast, with the average lifetime of a CH3Hg(II)-glutathione complex estimated to be less than 0.01 s. From exchange-averaged chemical shifts of the resonance for the proton on the alpha-carbon of the cysteine residue of glutathione, it is shown that, in hemolyzed erythrocytes, the sulfhydryl group of glutathione binds CH3Hg(II) more strongly than the sulfhydryl groups of hemoglobin.

1H nmr study of the effectiveness of various thiols for removal of methylmercury from hemolyzed erythrocytes

The effectiveness of eight thiol ligands for removing methylmercury (CH3Hg(II)) from its glutathione and hemoglobin complexes in hemolyzed erythrocytes has been studied by 1H nuclear magnetic resonance spectroscopy. These complexes are the predominant methylmercury species in human erythrocytes. The effectiveness was determined from the exchange-averaged chemical shift of the resonance for the proton on the alpha-carbon of the cysteinyl residue and from the intensity of the resonance for the methylene protons of the glycine residue of reduced glutathione (GSH), both of which provide a measure of the amount of glutathione in the CH3Hg(II)-complexed form. The thiol ligands were found to release GSH from its CH3Hg(II) complex in the order 2, 3-dimercaptosuccinic acid greater than mercaptosuccinic acid greater than cysteine greater than mercaptoacetic acid greater than D-penicillamine greater than 2,3-dimercaptopropanesulfonic acid greater than N-acetyl-D,L-penicillamine greater than D,L-homocysteine.