The redox reactivities of air-oxidized apo horse spleen ferritin (HoSF) and apo rat liver ferritin (RaF) were examined by microcoulometry and reductive optical titrations. Microcoulometry on several independent lots of commercial HoSF revealed two distinct types of redox activity: one requiring 3-4 electrons and one requiring 6-7 electrons for full reduction of the protein shell. ApoRaF required 8-9 electrons to fully reduce the oxidized form. Reductive optical titrations confirmed the microcoulometric reduction stoichiometry and, in addition, showed that the spectra of both oxidized and reduced apoHoSF were distinct and possessed absorbances tailing into the visible region. The redox reactivity of both apoRaF and apoHoSF correlated with their H-subunit composition. Identical microcoulometric and optical experiments were conducted with recombinant apo human liver heavy (rHuHF) and light (rHuLF) ferritins, but neither was redox-active. These results suggest that the redox reactivity of native ferritins is due to their heteropolymeric nature. This was confirmed by mixing various proportions of rHuHF and rHuLF, dissociating the 24-mers into individual subunits with guanidine hydrochloride at pH 3.5, and renaturing to form heteropolymeric 24-mers. Microcoulometric measurements of these apoheteropolymers reassembled in vitro showed that they were redox-active like their native apoheteropolymer counterparts. The redox activity of these apoheteropolymers increased with H- subunit composition, reached a maximum near 12 H- and 12 L-subunits, and then declined to zero with increasing L-subunit composition. The decline in redox reactivity at high L-subunit concentrations indicates that both H- and L- subunits are involved in forming the observed redox centers. Apoheteropolymers formed from rHuLF and W93F (an H-chain mutant) were redox- inactive, suggesting that the conserved tryptophan is necessary for redox center formation.