The iron-storage protein bacterioferritin (BFR) from Escherichia coli consists of twenty four identical subunits, each containing a dinuclear metal ion-binding site (the ferroxidase centre) at which iron(II) is oxidised to iron(III) and dioxygen is reduced. Other metal ions that are commonly found in biological systems bind to the ferroxidase centre, including manganese(II), cobalt(II) and zinc(II). In this work, copper(II)-binding to BFR and its effect on iron(II) oxidation kinetics were studied by a combination of gel filtration–copper(II) binding assay, optical, magnetic and kinetic methods. Data indicate that two copper(II) ions bind per subunit with a K_d of ≈ 2.0 × 10⁻⁵ M and establish the order of divalent metal ion binding as Cu(II) < Co(II) < Zn(II), i.e. it does not follow the Irving–Williams order. A number of lower affinity copper(II)-binding sites were also detected. The presence of copper(II) was found to significantly enhance the rate of iron(II) oxidation and subsequent core formation. This effect does not arise from copper(II) bound at the ferroxidase centre but, rather, is due to displaced copper(II). The nature of the displaced copper is discussed.