Acridones (9-azaanthracen-10-ones), especially halogen-substituted acridone-carboxylic acids are efficient inhibitors of [14C]ADP binding to the bovine mitochondrial ADP/ATP-carrier protein. 7-Iodoacridone-4-carboxylic acid displaces [14C]ADP in a competitive manner, which indicates that both compounds share an identical binding site. Upon ultraviolet illumination, 7-azido-4-[1',2'-3H]isopropyl-acridone binds covalently to the ADP/ATP-carrier protein. By sequencing of its proteolytic fragments, radioactivity was detected in Cys159. This amino acid is located close to Lys162, a target of 2-azido-[alpha-32P]ADP [Dalbon, P., Brandolin, G., Boulay, F., Hoppe, J. & Vignais, P. V. (1988) Biochemistry 27, 5141-5149].