Abstarct: The polygalacturonase of Macrophomina phaseolina, causal agent of charcoal rot disease in wide range of plants, seems to be involved in peneteration into the host plant cell wall. Ion exchange chromatography was used to isolate the enzyme. The enzyme had a relative molecular mass of 80 kDa. It’s optimum activity was at pH = 3 and 60 ⁰C, being stable at wide ranges of pH and temperature (t1/2 of 177 min at 90°C), making it useful for industrial applications. The activation energy and activation enthalpy for heat inactivation was 15.26 kJ/mol, and 12.41 kJ/mol, respectively. Kinetics of enzyme analysis (nH = 10.1, Rs = 1.7) revealed a non-typical Michaelis–Menten pattern, illustrating a sigmoidal curve with Vmax = 0.008 µmol/min and K0.5 = 3 mg/ml. The data was indicative of the presence of at least two catalytic sites with positive allosteric interaction between polygalacturonase and the substrate (Polygalacturonic acid). Triton X-100, MnCl2, FeSO4, MnSO4 and CoCl2 stimulated polygalacturonase activity, whereas Tween-20, Tween-80, PMSF, NiSO4, BaCl2, CuSO4, LiCl and phosphate salts inhibited the enzyme activity. Iodoacetamide and iodoacetic acid did not inhibit enzyme activity, suggesting that cysteine residues are not part of the catalytic site of polygalacturonase.