Nuclear Magnetic Relaxation Dispersion in Protein Solutions

To investigate the role of specifically bound bicarbonate in the metal-complexing function of transferrin, copper transferrin was prepared both in the presence and absence of bicarbonate ion. The magnetic field and temperature dependence of the solvent proton spin-lattice relaxation rate of solutions of the two complexes were determined. The molar relaxivity R (essentially, relaxation rate per protein concentration) for solutions of copper transferrin is very large at low fields (∼20,000 m-1 sec-1 at 4 Oe) and is highly field-dependent, decreasing to almost zero at 12 kOe. At low field, R is almost temperature-independent over the range 0–45°. Comparison of the results with theory indicates that protons are relaxed by a dipolar interaction with the unpaired electronic spin of Cu2+. The observed relaxivity results from the rapid exchange of a water molecule bound sufficiently close to the copper to be an axial ligand. The magnitude of R at low field for copper transferrin-HCO3 is about one-quarter of that for copper transferrin and also disperses almost to zero at high field. Above 25° the low field value of R has a strong positive temperature dependence. There is no significant variation in R over the pH range 7 to 9.5. The data suggest that, as in the absence of bicarbonate, the protons of water molecules are relaxed by a dipolar interaction with the electron spin of Cu2+, and that bicarbonate binding alters the water exchange rate.