Differential Interactions of the Mg²⁺ Complexes of Chromomycin A₃ and Mithramycin with Poly(dG-dC)·Poly(dC-dG) and Poly(dG)·Poly(dC)

Abstract

The interaction of the two anticancer antibiotics, chromomycin A3 and mithramycin, with the polynucleotides poly(dG-dC) x poly(dC-dG), representative of B-DNA, and poly(dG) x poly(dC), representative of A-DNA, in the presence of Mg2+ is studied by spectroscopic techniques such as absorbance, fluorescence, and dircular dichroism (CD). The studies were done with both drug x Mg2+ complexes, I and II, having 1:1 and 2:1 stoichiometries with respect to drug and Mg2+, respectively [Aich, P., Sen, R., & Dasgupta, D. (1992) Biochemistry 31, 2988-2997]. The objective of the present work is 2-fold. First, an attempt is made to understand the structural basis of the ligand-DNA interaction, particularly the role of DNA backbone conformation with its groove size and the accessibility of the 2-amino group in the minor groove of guanosine. Second, the role of the antibiotic saccharide moieties in the association with DNA was studied. For this purpose, the spectroscopic characterization of the binding was done followed by the evaluation of binding parameters and associated thermodynamics. Analysis of the observed thermodynamics for the ligand-DNA interactions in terms of the different structures of the polynucleotides was done. The salient results are as follows. Complex I does not discriminate significantly among the A- and B-forms of DNA when it binds to them in an entropy-driven process. On the other hand, complex II for both drugs recognizes B- and A-forms of DNA in different ways. This observation implies that the sequence specificity shown by this complex is a sequel to the difference in the parameters such as groove size and accessibility of the guanosine amino group. Another important finding is that binding with the same polynucleotide is not comparable for the complex II of the two drugs. It emphasizes the involvement of the sugar moieties, when the drug x Mg2+ complex binds to DNA. The presence of an acetoxy group in the sugars of chromomycin A3 imparts some distinctive specific features of the association of the chromomycin dimer x Mg2+ complex with DNA. Finally, the results are compared with those available from NMR studies of different drug-oligonucleotide complexes under conditions where complex II is the ligand.

Structural basis of DNA recognition by anticancer antibiotics, chromomycin A3, and mithramycin: Roles of minor groove width and ligand flexibility

“…Stoichiometry of binding was also determined from the point of intersection of the two straight lines in the plot of normalized increase in fluorescence against the ratio of the input concentrations of oligomer and ligand. 12 In every case stoichiometry was obtained as number of bound nucleotides per drug molecule.…”

“…Spectrofluoremetric titration was also used to determine binding stoichiometry from the intersection of the straight lines in the plot of normalized increase in fluorescence intensity as a function of input concentration of oligomer and drug ratio. 12 A representative example for association of (MTR) 2 Mg 2ϩ complex with d(CCGGCGCCGG) 2 is shown in Figure 5b. The binding stoichiometry comes out to be same within the limits of experimental error, by both methods of evaluation.…”

“…15,16 Structural studies on the association of CHR and MTR with DNA have also implicated the importance of groove dimension during recognition process. 12,17,18 Analysis of thermodynamic parameters associated with the antibiotic-DNA recognition indicated a structural transition of the DNA at the binding site. 12 NMR studies also suggested a localized B 3 A transition at the binding site for association of complex II with B-DNA.…”

“…12,17,18 Analysis of thermodynamic parameters associated with the antibiotic-DNA recognition indicated a structural transition of the DNA at the binding site. 12 NMR studies also suggested a localized B 3 A transition at the binding site for association of complex II with B-DNA. 18,19 These studies showed that H bonding between ONH2 proton of guanosine base with the OOH group at 8 position in the aglycone is the origin of GC base specificity for the antibiotics.…”

“…In an earlier report, we showed that alteration in DNA backbone geometry affects accessibility of the potential H-bonding sites in DNA minor groove. 12 B 3 A transition at the binding site containing GC-rich sequence leads to widening of the minor groove, as a result of which a higher number of NH2 protons of guanosine base gets exposed for H bonding with the ligand(s). In a separate study, we have compared the conformational flexibility of complex II for the two antibiotics during its interaction with DNA.…”