Interactions of iron bleomycin, phosphate or cyanide, and DNA: sequence-dependent conformations and reactions.
The hypothesis was investigated that axial ligands bound to Fe(III)-bleomycin [Fe(III)Blm] are destabilized at specific 5'-guanine-pyrimidine-3' binding sites but are stable at nonselective dinucleotides. DNA oligomers and calf-thymus DNA were used in reactions with L-Fe(III)Blm, where phosphate and cyanide served as examples of large and small ligands (L). Both ligands underwent dissociation when L-Fe(III)Blm was bound to d(GGAAGCTTCC)2 (I) but not d(GGAAATTTCCC)2 (II) and at large ratios of calf-thymus DNA to drug. Fe(III)Blm is high spin in 20 mM phosphate buffer, signifying the presence of a phosphate adduct. In the titration of HPO4-Fe(III)Blm with calf-thymus DNA, a large excess of DNA was needed to reach the low-spin state, consistent with an equilibrium competition between phosphate and DNA for Fe(III)Blm. Equilibrium constants for binding Fe(III)Blm and CN-Fe(III)Blm to calf-thymus DNA (6.8x10(5) M(-1) and 5.9x10(4) M(-1), respectively, in HEPES buffer at 25 degrees C and pH 7.4) showed that the CN- ligand also reduced the affinity of DNA for the drug. The kinetics of dissociation of CN- from CN-Fe(III)Blm-DNA were slow and first order in bound drug. The reversible nature of these dissociation reactions was shown using 1H NMR spectroscopy of Fe(III)Blm-I in the absence and presence of large excesses of CN- or phosphate. The results are discussed in terms of a two-state hypothesis for the binding of L-Fe(III)Blm to specific and nonspecific dinucleotides. It is proposed that steric restrictions at specific sites inhibit binding of these ligands.
Li
W
W
Department of Chemistry, University of Wisconsin-Milwaukee, 53201, USA.
Xia
C
C
Antholine
W E
WE
Petering
D H
DH
D020029:Base Pairing; D001761:Bleomycin / Q000737:chemistry / Q000378:metabolism*; D003486:Cyanides / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D018736:DNA Adducts / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D007501:Iron / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008968:Molecular Conformation; D009690:Nucleic Acid Conformation; D010710:Phosphates / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D003486:Cyanides; D018736:DNA Adducts; D010710:Phosphates; D001761:Bleomycin; D004247:DNA; D007501:Iron