A unique binding cavity for divalent cations in the DNA-metal-chromomycin A3 complex

Binding of chromomycin A3 (CRA) to calf thymus DNA was investigated in the presence of divalent cations using visible absorption and 1H-nmr spectroscopies. An apparent equilibrium binding constant (approximately 10(11) M-1) was obtained from metal competition experiments using EDTA to remove the metal cation from the DNA-M-CRA (M: metal) complex. The large binding constant of the drug to DNA enabled us to obtain essentially complete complexation of CRA to the short homogeneous d(ATGCAT)2 duplex using stoichiometric amounts of the metal cation. Large induced chemical shifts were observed in the 1H-nmr spectrum of the above complex using the paramagnetic Co2+ cation, indicating that the metal occupies a unique binding site. Since no induced 1H-nmr chemical shifts were observed for the drug-Co2+ mixture, it was concluded that no metal-drug complex is formed. In addition, it was found that bound CRA is negatively charged at physiological pH and binding to the DNA could be affected only by using metal cations whose ionic radius size (less than 0.85 A) and charge (2+) were simultaneously satisfied. Stringent metal cation selectivity for the DNA-M-CRA complex may be intimately connected with the antitumor selectivity of CRA, since different types of cells generally possess widely differing molar concentrations of metal cations.