Isolation of a beta-galactosidase from chicken liver.
In search of a beta-galactosidase which specifically hydrolyses beta 1----3 bound galactose residues in galacto-glycoconjugates, an acid beta-galactosidase from chicken liver was investigated. The isolation procedure involved ammonium sulphate precipitation followed by lectin chromatography on Con A-Sepharose 4B, ion-exchange chromatography on DEAE-cellulose, gel filtration on Sepharose 6B and affinity chromatography on p-aminophenyl-thio-beta-D-galactoside-agarose. The beta-galactosidase was purified 3000-fold with 11% recovery of enzyme activity. The purified protein showed an apparent molecular mass of above 200000 in SDS-polyacrylamide gel electrophoresis. A few minor bands were also present. The reduced and denatured beta-galactosidase migrated as a single major band with an apparent molecular mass of 67000. The enzyme released galactose from lactose and from the synthetic substrates Gal beta 1----3Gal, Gal beta 1----6Gal and Gal beta 1----3Ara. However, the enzyme did not release galactose from the snail gland galactans and the high molecular weight galacto-glycoconjugates and it did not hydrolyse the peanut agglutinin receptor of the red blood cell membrane.
D000645:Ammonium Sulfate; D000818:Animals; D002384:Catalysis; D002645:Chickens; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D004591:Electrophoresis, Polyacrylamide Gel; D005696:Galactosidases / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology*; D012967:Sodium Dodecyl Sulfate; D013379:Substrate Specificity; D001616:beta-Galactosidase / Q000302:isolation & purification* / Q000378:metabolism
D012967:Sodium Dodecyl Sulfate; D005696:Galactosidases; D001616:beta-Galactosidase; D000645:Ammonium Sulfate