Purification and characterization of sialidase from porcine liver

Abstract

Sialidase [E.C.3.2.1.18] has previously been purified from porcine liver by procedures including extraction, ammonium sulfate precipitation, concanavalin A-Sepharose adsorption, activation, CM-Sepharose ion exchange chromatography, and HPLC on a Shim pack Diol 300 column. Two sialidase preparations, sialidase I and II, were obtained by CM-Sepharose column chromatography and were eluted with pH 4.5 and 5.0 buffers, respectively. The two enzyme preparations showed the same optimum pH, pH stability, and specificities for natural substrates. The two final preparations contained β-galactosidase activity and showed three protein components of 64, 30, and 21 kDa with sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which are derived from the β-galactosidase multimer. The anti-β-galactosidase multimer antiserum was able to precipitate sialidase activity. It is likely that porcine liver sialidase exists as a multienzyme complex with β- galactosidase and carboxypeptidase (protective protein).