Purification and characterization of ovine pancreatic elastase

Elastase was isolated from ovine pancreas and purified to homogeneity by two different procedures. One involved precipitation with ammonium sulphate, p-aminobenzamidine–Sepharose chromatography, CM-Sepharose ion exchange chromatography and S-300 Sephacryl chromatography. The other involved the direct adsorption of elastase by tri- l-alanyl–Sepharose chromatography and a CM-Sepharose step. The enzyme, which was produced in an inactive form in the pancreas, was activated with a trace of trypsin prior to chromatography. Ovine pancreatic elastase has an isoelectric point above p I 9.3 and its molecular mass is estimated at ∼25 kDa. The optimal pH range for activity is between 8.0 and 8.4 and the enzyme is unstable at pH below 4.0 and above 10.0 and at temperatures above 65°C. The kinetic properties of the enzyme were determined with succinyl-Ala-Ala-Ala- p-nitroanilide as the substrate. K m and k cat K m −1 proved to be similar to the kinetic parameters of porcine elastase determined simultaneously.