The stoichiometry and stability of the NADP complexes with manganese(II) ions as studied by electron paramagnetic resonance

Magnetic resonance techniques have been applied to study the stability of the complexes formed between Mn(II) ions and NADP in aqueous solutions at a pH of 7.5 and 20 °C. The electron paramagnetic resonance (epr) data indicate that at low Mn(II) ion concentrations ([Mn(II)] < 1 mM; [NADP] ~ 5 mM), a 1:1 complex is formed with an apparent stability constant K1 = 370 ± 50 M−1 at an ionic strength of 0.22 in the presence of 0.20 M Cl−. At high Mn(II) ion concentrations, a Mn(II)2 –NADP species, with an apparent stability constant K2 = 54 ± 17 M−1, is present in significant amounts. When the epr data are corrected for the presence of the MnCl+ ion, the analysis of the new Scatchard plot yields stability constants for the two sites of K1 = 640 ± 90 M−1 and K2 = 88 ± 13 M−1 respectively. The presence of two metal ion binding sites on the NADP molecule has not been observed previously, and previous workers have always analyzed their data in terms of the 1:1 Mn(II)–NADP complex. An epr temperature study of K1 yields a value of ΔH equal to 1.3 ± 0.2 kcal/mol (1 cal = 4.187 J).