Department of Biochemistry, Faculty of Science, Saitama University, 255 Shimo-okubo, Urawa 338, Japan.
Plant Physiol. 1989 Jun;90(2):567-74. doi: 10.1104/pp.90.2.567.
A basic beta-galactosidase (beta-Galase) has been purified 281-fold from imbibed radish (Raphanus sativus L.) seeds by conventional purification procedures. The purified enzyme is an electrophoretically homogeneous protein consisting of a single polypeptide with an apparent molecular mass of 45 kilodaltons and pl values of 8.6 to 8.8. The enzyme was maximally active at pH 4.0 on p-nitrophenyl beta-d-galactoside and beta-1,3-linked galactobiose. The enzyme activity was inhibited strongly by Hg(2+) and 4-chloromercuribenzoate. d-Galactono-(1-->4)-lactone and d-galactal acted as potent competitive inhibitors. Using galactooligosaccharides differing in the types of linkage as the substrates, it was demonstrated that radish seed beta-Galase specifically split off beta-1,3- and beta-1,6-linked d-galactosyl residues from the nonreducing ends, and their rates of hydrolysis increased with increasing chain lengths. Radish seed and leaf arabino-3,6-galactan-proteins were resistant to the beta-galase alone but could be partially degraded by the enzyme after the treatment with a fungal alpha-l-arabinofuranosidase leaving some oligosaccharides consisting of d-galactose, uronic acid, l-arabinose, and other minor sugar components besides d-galactose as the main product.
已通过常规提纯程序从吸水萝卜（Raphanus sativus L.）种子中提纯出 281 倍的基础β-半乳糖苷酶（β-Galase）。纯化后的酶是一种电泳均一的蛋白质，由一条单多肽组成，其表观分子量为 45 千道尔顿，pl 值为 8.6 至 8.8。该酶在 pH4.0 时对 p-硝基苯-β-d-半乳糖苷和β-1,3-连接的半乳糖二糖的活性最大。酶的活性被 Hg（2+）和 4-氯汞苯甲酸强烈抑制。d-半乳糖酸-（1->4）-内酯和 d-半乳糖醛酸是有效的竞争性抑制剂。用作为底物的不同键型的半乳糖低聚糖进行实验，证明萝卜种子β-Galase 特异性地从非还原端切割β-1,3-和β-1,6-连接的 d-半乳糖基残基，其水解速率随链长的增加而增加。萝卜种子和叶阿拉伯-3,6-半乳聚糖蛋白对β-半乳糖苷酶本身具有抗性，但在用真菌α-l-阿拉伯呋喃糖苷酶处理后，该酶可部分降解，留下一些由 d-半乳糖、糖醛酸、l-阿拉伯糖和除 d-半乳糖以外的其他少量糖成分组成的低聚糖作为主要产物。
