PURIFICATION AND SOME PROPERTIES OF A CARBOXYLESTERASE FROM OVINE LIVER

Carboxylesterase ESB3 was extracted from ovine liver and purified to h omogeneity by ammonium sulphate fractionation, hydrophobic interaction chromatography on Phenyl Sepharose, ion exchange chromatography on Mo no-Q Sepharose and size exclusion chromatography on Superose 6. The en zyme is free of carboxylesterase ESB2 activity. The molecular mass of the enzyme is estimated 182 kDa as judged by size exclusion chromatogr aphy. Isoelectric focusing indicates the presence of six isoforms of p I 5.50-5.77 with three main isoforms of pI 5.55-5.65. The enzyme is ac tive towards the substrates p-nitrophenyl acetate and the aliphatic su bstrates ethyl acetate, ethyl propionate, ethyl butyrate, and ethyl va lerate. Of the ethyl esters the affinity is lowest towards acetate and highest towards ethyl butyrate. The enzyme is totally inhibited by ph enylmethylsulphonyl fluoride (PMSF) and mercuric chloride but not affe cted by eserine or cupric chloride. The pH optimum of the enzyme is 7. 5 and it is stable at 55 degrees C for 20 min.