Javeri S, Uhlenbruck G
J Clin Chem Clin Biochem. 1984 Nov;22(11):735-9. doi: 10.1515/cclm.1984.22.11.735.
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.
为了寻找一种能特异性水解半乳糖糖缀合物中β-1,3连接的半乳糖残基的β-半乳糖苷酶，对鸡肝中的一种酸性β-半乳糖苷酶进行了研究。分离过程包括硫酸铵沉淀，随后在Con A-Sepharose 4B上进行凝集素色谱、在DEAE-纤维素上进行离子交换色谱、在Sepharose 6B上进行凝胶过滤以及在对氨基苯基硫代-β-D-半乳糖苷-琼脂糖上进行亲和色谱。β-半乳糖苷酶被纯化了3000倍，酶活性回收率为11%。纯化后的蛋白质在SDS-聚丙烯酰胺凝胶电泳中显示表观分子量高于200000。还存在一些次要条带。还原和变性后的β-半乳糖苷酶以单一主要条带形式迁移，表观分子量为67000。该酶能从乳糖以及合成底物β-1,3连接的半乳糖、β-1,6连接的半乳糖和β-1,3连接的阿拉伯糖中释放半乳糖。然而，该酶不能从蜗牛腺半乳聚糖和高分子量半乳糖糖缀合物中释放半乳糖，也不能水解红细胞膜上的花生凝集素受体。
