31P NMR lineshapes of β-P (ATP) in the presence of mg2+ and ca2+ : estimate of exchange rates

The ³¹ P NMR chemical shift of β-P of adenosine triphosphate (ATP) undergoes a substantial change (2̃–3 ppm) upon chelation of divalent ions such as Mg²⁺ or Ca²⁺. In the presence of nonsaturating amounts of Mg²⁺ or Ca²⁺, the lineshape of this resonance depends on the characteristic association and dissociation rates of these metal-ATP complexes. A procedure for computer simulation of this lineshape is outlined. A comparison of computer-simulated lineshapes with the experimental lineshapes obtained at 121 MHz was used to determine the following dissociation rate of Mg²⁺ and Ca²⁺ from their ATP complexes at 20°C and pH 8.0: Ca²⁺, > 3 × 10⁵ s^?1 (Hepes buffer); Mg²⁺, 1200 s^-1 (no buffer), 1000 s^-1 (Tris buffer) and 2100 s^?1 (Hepes buffer). The limits of error are ± 10% in these values. For the Mg²⁺ complexes, the rates were determined as a function of temperature to obtain activation energies (with a maximum deviation of 10% in the least-squares fit): 8.1 $\text{Kcal}\text{mole}$ (no buffer and Hepes buffer) and 6.8 $\text{kcal}\text{mole}$ (Tris buffer). Lineshapes of the β-Presonance simulated as a function of Mg²⁺ concentration, using 2100 s^?1 for the dissociation rate, are also presented. The computer simulation of lineshapes offers a reliable and straightforward method for the determination of exchange rates of diamagnetic cations from their ATP complexes, under a variety of sample conditions.