The Mechanism of Water Proton Nuclear Magnetic Resonance Relaxation in the Presence of Mammalian and Aplysia Metmyoglobin Fluoride

A mechanism of proton exchange leading to water proton relaxation is proposed for horse heart metmyoglobin fluoride which is mediated by H2O molecules and involves the distal histidine. The mechanism is based on a comparison of water proton NMR relaxation (T1-1 (H)) in the presence of horse and Aplysia metmyoglobin fluoride; Aplysia metmyoglobin fluoride was chosen for comparison because it lacks the distal histidine. Proton relaxation for horse heart metmyoglobin fluoride has been observed over a pH range of 5.0 to 11.5 in H2O and 95% D2O solutions while simultaneously monitoring the optical absorbance at 542 nm. For horse heart metmyoglobin fluoride, proton relaxation shows three pK values, at pH 6.2, 8, and one at a higher pH, which varies with the fluoride ion concentration due to the equilibrium metmyoglobin fluoride + OH- ⇋ MbOH + F-. Optical spectroscopy shows two pK values, one at pH 6.2 and one above 9. The latter pK has a large deuterium isotope effect in the optical spectrum, but none for proton relaxation. Theorell and Ehrenberg ((1951) Acta Chem. Scand. 5, 823) reported changes in the magnetic susceptibility of metmyoglobin fluoride with pK values of 6.3 and 8.11; however, the magnetic susceptibility increases when T1-1 (H) decreases and vice versa. We therefore conclude that the proton-iron distance changes twice during the titration.