SPECTROSCOPIC STUDIES OF COBALT(II) BINDING TO ESCHERICHIA-COLI BACTERIOFERRITIN

The iron storage protein bacterioferritin (BFR) consists of 24 identic al subunits, each containing a dinuclear metal binding site called the ferroxidase center, which is essential for fast iron core formation, Cobalt(II) binding to wild-type and site-directed variants of Escheric hia coli BFR was studied by optical and magnetic techniques, Data from absorption spectroscopy demonstrate the binding of two cobalt(II) ion s per subunit of wild type and heme-free BFR, each with a pseudotetrah edral or pentacoordinate geometry, and EPR studies show that the two c obalt(II) ions are weakly magnetically coupled. Studies of variants of BFR in which a single glutamic acid residue at the ferroxidase center is replaced by alanine confirm that this is the site of cobalt(II) bi nding, since the altered centers bind only one cobalt(II) ion, This wo rk shows that the electroneutrality of the ferroxidase center is prese rved on binding a pair of divalent metal ions. Optical and EPR data sh ow that cobalt(II) binding to BFR exhibits positive cooperativity, wit h an average K-d of similar to 1 x 10(-5) M. The favored filling of th e ferroxidase center with pairs of metal ions may have mechanistic imp lications for the iron(II) binding process, Discrimination against oxi dation of single iron(II) ions avoids odd electron reduction products of oxygen.