N-lobe versus C-lobe complexation of bismuth by human transferrin

Interactions of recombinant N-lobe of human serum transferrin (hTF/2N) with Bi³⁺, a metal ion widely used in medicine, have been investigated by both UV and NMR spectroscopy. The bicarbonate-independent stability constant for Bi³⁺ binding (K*) to hTF/2N was determined to be log K* 18.9±0.2 in 5 mM bicarbonate/10 mM Hepes buffer at 310 K, pH 7.4. The presence of Fe³⁺ in the C-lobe of intact hTF perturbed Bi³⁺ binding to the N-lobe, whereas binding of Bi³⁺ to the C-lobe was unaffected by the presence of Fe³⁺ in the N-lobe. Reactions of Bi³⁺ (as bismuth nitrilotriacetate or ranitidine bismuth citrate) with hTF/2N in solutions containing 10 mM bicarbonate induced specific changes to high-field ¹H-NMR peaks. The ¹H co-ordination shifts induced by Bi³⁺ were similar to those induced by Fe³⁺ and Ga³⁺, suggesting that Bi³⁺ binding causes similar structural changes to those induced by hTF/2N. ¹³C-NMR data showed that carbonate binds to hTF/2N concomitantly with Bi³⁺.