MG-25 NMR-STUDIES OF MAGNESIUM BINDING TO ERYTHROCYTE CONSTITUENTS

The binding of Mg2+ ion to ATP, ADP, AMP, 2,3-bisphosphyoglycerate (DPG), and hemoglobin has been studied by Mg-25 NMR spectroscopy at 9.4 T. Addition of any of these ligands to a solution of 2 mM (MgCl2)-Mg-25 at pH 7.2 caused a progressive increase in linewidth, with no discernible chemical shift. ATP and ADP, which form tight 1:1 complexes with Mg2+, did not cause maximal broadening until present in several-fold excess, implying that bis(nucleotide) complexes also form. The studies showed progressively weaker Mg2+ binding to ATP, ADP, DPG, and AMP, consistent with published binding constants. Hemoglobin cause fairly little broadening, consistent with its known weak affinity for Mg2+. Competition studies determined ATP affinities for Ca2+ and H+ that were also in good agreement with published values. Mg-25 NMR spectra of 2 mM bound Mg-25(2+) were obtained with good signal to noise in less than 1 hr. The technique may now be a practical means for studying the binding of Mg2+ within erythrocytes and other cells.