In agreement with previous work [Shuvalov, Heber and Schreiber (1988) FEBS Lett. 258, 27-31] two hemes (low potential (LP) and extra low potential (XLP )) per two pheophytins were found in isolated D1D2Cyt b-559 complexes. Reductive and oxidative redox titrations demonstrate that the E m of the LP form is at about +150 mV. It is independent of pH between pH 7.2 and 9.4. The XLP heme is autoxidizable at pH 7.2 and displays, at this pH, an E m of − 45 mV. Both the LP and XLP hemes show absorption peaks at 559 nm. They are proposed to have bis-histidine ligation of the heme iron. At pH 9.4, the XLP heme splits into two forms. One of them has an E m of +40 mV, and absorption peaks at 559 nm showing the bis-histidine ligation. The other displays an E m of −220 mV and the peak is shifted to 562 nm. This last form is proposed to be due to the incorporation of OH − which occupies the 6th coordination position of the heme Fe(III) at high pH. The p K value for the conversion of the XLP heme is close to 7.7. In a structure simulation of the α-helices of α- and β-polypeptides, the β-polypeptide, but not the α-polypeptide, reveals a distance between the histidine N and the heme Fe which permits stable N-Fe coordination. In the α-polypeptide, OH − can be incorporated between N and Fe. The functional role of the two hemes of cyt b-559 is briefly discussed with respect to water oxidation and cyclic electron transfer.