Nuclear Magnetic Resonance Study of Metal-Ion Binding to Adenosine Triphosphate. I. 31P Studies

Measurements of the 31P NMR of the α, β, and γ phosphates of adenosine triphosphate (ATP) have been made for Mn2+—, Ni2+—, Co2+—, and Cu2+—ATP complexes. In order to explain the metal-ion—phosphate interaction, measurements were made at different values of f≡M2+/ATP between 0° and 95°C. The metal ions broadened the 31P NMR, described by a decrease in the observed value of T2; reduced T1, the spin—lattice relaxation time; and in the case of CoATP at high temperatures, shifted the resonance to lower fields by an amount ΔωM because of the transferred isotropic hyperfine coupling A. For Mn2+ and Ni2+ at all temperatures and CoATP at lower temperatures, T2 was determined by the chemical exchange time τM of the metal—phosphate bonds, and all three phosphates had the same widths. For CoATP at higher temperatures, the resonances were exchange narrowed and 1/T2≈fτMΔωM2. In agreement with the shift measurements 1/T2|γ1/T2|β1/T2|α. The values of A from the shifts were (in megacycles per second): Aγ=5.4, Aβ=4.5, and Aα=3.6 which were slightly larger than the values in Co3(PO4)2 crystals of 3.3. The large values of A in ATP show that the Co2+ is bound all the time to the phosphates. The T1 values for MnATP complexes are determined by dipolar interactions of the form 1/T1 ∝ (μ/r3)2τc. At room temperatures T1|γ=T1|β=34T1|α, which implies a very small difference in the distance of the metal ion from all three 31P nuclei. At higher temperature all three were equal. Assuming the phosphorous—metal distance of 3.3 Å, τc is determined to be ∼1×10−9 sec, which is about four times larger than the same correlation time for the water protons. From the high-temperature MnATP linewidths, which were all equal, we conclude that the hyperfine coupling with all three 31P nuclei is 0.6 times the value of A observed by Mays in LiMnPO4 crystals, once again showing that the binding to all three phosphates is approximately complete and approximately the same.