Thermodynamics and kinetics of the cleavage of DNA catalyzed by bleomycin A5 -: A microcalorimetric study

Microcalorimetry and UV-vis spectroscopy were used to conduct thermodynamic and kinetic investigations of the scission of calf thymus DNA catalyzed by bleomycin A(5) (BLM-A(5) ) in the presence of ferrous ion and oxygen. The molar reaction enthalpy for the cleavage, the Michaelis-Menten constant for calf thymus DNA and the turnover number of BLM-A(5) were calculated by a novel thermokinetic method for an enzyme-catalyzed reaction to be -577 +/- 19 kJ.mol(-1) , 20.4 +/- 3.8 mum and 2.28 +/- 0.49 x 10(-2) s(-1) , respectively, at 37.0 degreesC. This DNA cleavage was a largely exothermic reaction. The catalytic efficiency of BLM-A(5) is of the same order of magnitude as that of lysozyme but several orders of magnitude lower than those of Taq I restriction endonuclease, Nae I endonuclease and Bam HI endonuclease. By comparing the molar enthalpy change for the cleavage of calf thymus DNA induced by BLM-A(5) with those for the scission of calf thymus DNA mediated by adriamycin and by (1,10-phenanthroline)-copper, it was found that BLM-A(5) possessed the highest DNA cleavage efficiency among these DNA-damaging agents. These results suggest that BLM-A(5) is not as efficient as a DNA-cleaving enzyme although the cleavage of DNA by BLM-A(5) follows Michaelis-Menten kinetics. Binding of BLM-A(5) to calf thymus DNA is driven by a favorable entropy increase with a less favorable enthalpy decrease, in line with a partial intercalation mode involved in BLM-catalyzed breakage of DNA.