13C Nuclear Magnetic Resonance Study of the Spatial Relation of the Metal- and Anion-binding Sites of Human Transferrin

Abstract One carbonate anion of the diamagnetic Co3+-transferrin-CO3 complex, which contains two metal ions and two carbonate anions, exchanges rapidly with free bicarbonate in solution while the other does not exchange at an appreciable rate. The 13C nuclear magnetic resonance absorption of the nonexchanging carbonate has been observed as a line, 14 Hz wide, occurring 104 ppm downfield of 1,4-dioxane and 8.0 ppm downfield of free bicarbonate in the same solution. The analogous absorption of the paramagnetic Fe3+-transferrin-13CO3 complex is broadened beyond detection. This broadening places an upper limit of about 9 A on the Fe3+-13C distance but is also consistent with smaller values, such as might result from direct binding of carbonate to iron. Both carbonate ions of the specific transferrin complexes of Ga3+, Zn2+, and Cu2+ exchange in minutes with bicarbonate in solution, making preparation of 13C-enriched complexes difficult.