Oxygen-dependent Oxidation of Fe(II) to Fe(III) and Interaction of Fe(III) with Bovine Serum Albumin, Leading to a Hysteretic Effect on the Fluorescence of Bovine Serum Albumin

The serum albumin is the most abundant protein in blood plasma and the iron is essential for many cellular processes. However, the interaction between Fe3+ and haem-free serum albumin remains unclear. Here we provide evidence for the fact that haem-free BSA possesses one specific Fe3+-binding site. The binding of Fe3+ to BSA results in a significant quenching of the Trp fluorescence of BSA. The average apparent dissociation constant value for the interaction of Fe3+ and BSA is 3.46 × 10−8 ± 3 × 10−10 M at 37 °C and 3.30 × 10−8 ± 5 × 10−10 M at 25 °C, respectively, as determined by fluorescence titration. Addition of 50 μM Fe2+ to 1 μM BSA results in an obvious hysteretic effect on the fluorescence of BSA. The time-dependent fluorescence quenching of BSA by Fe2+ is not caused by the Fe2+-induced conformational change of BSA, but the oxygen-dependent oxidation of Fe2+ to Fe3+. Fe2+ undergoes an oxygen-dependent oxidation to Fe3+ under aerobic conditions, which is accelerated by the interaction of BSA with Fe3+ and extensively inhibited under anaerobic conditions. The results suggest that BSA may take part in non-transferrin bound iron transfer.