An NMR Investigation of the Kinetics of Dissociation of the Zinc(II) Complex of Bleomycin Antibiotics

The addition of Zn(II) to a neutral aqueous solution of bleomycin results in the observation of several new peaks in the H NMR spectrum concomitant with the reduction in intensity of several of the peaks observed in the metal-free antibiotic. This observation leads to the conclusion that for some of the protons in bleomycin the rate of dissociation of the Zn(Il)-bleomycin complex is much smaller than their chemical-shift difference in the free and complexed states. We have found that the Zn(II)-bleomycin complex has a 1:1 stoichiometry by measuring the relative intensity of the C4 resonance of the imidazole proton at various Zn(II) concentrations. Rates of dissociation of the complex were determined at temperatures between 303 and 353 K by means of NMR transfer of magnetization experiments. From an analysis of the temperature dependence of these rates we obtained values for the energy and entropy of activation of 16.2 kcal and 50 eu. Both the observed rates and the value of the energy of activation obtained in our analysis are more similar to parameters determined for Zn(II)-containing metal-loenzymes than for low molecular weight ligands. On this basis we suggest that in the Zn(II)-bleomycin complex the Zn(II) is more likely to be in a tetrahedral environment than the octahedral environment proposed in some models for the complex.