Purification and characterization of acetyl esterase from bull testes

Abstract

Acetyl esterase (acetic-ester acetylhydrolase, EC 3.1.1.6) from bull testes was purified 325-fold by ammonium sulphate precipitation, chromatography on DEAE-cellulose, hydroxyapatite and finally, gel filtration on a Sephadex G-200 column. The purified enzyme appeared as a single protein band on native polyacrylamide gel electrophoresis and in isoelectric focusing (p I 5.25). In both methods, the activity coincided with the protein band. A single protein band corresponding to M r 70 000 was obtained by SDS-polyacrylamide gel electrophoresis. The reported amino-acid composition indicates that the enzyme contains three half-cystine residues, of which only one could be detected, by titration, as a free -SH group. No free amino terminal was detected by dansylation.