Freilich L S, Lewis R G, Reppucci A C, Silbert J E
J Cell Biol. 1977 Mar;72(3):655-66. doi: 10.1083/jcb.72.3.655.
Suspension cultures of neoplastic mouse mast cells were used to obtain large quantities of a homogeneous cell population as starting material for cell fractionation. A Golgi fraction was prepared by slight modification of established techniques and identified by electron microscopy. Assay of galactosyl transferase activity using ovalbumin, desialylated degalactosylated orosomucoid, and N-acetylglucosamine as galactose acceptors showed that the Golgi fraction was enriched in specific activity over the homogenate. The Golgi galactosyl transferase was examined in detail. Acceptor concentrations for optimal galactose incorporation were determined, and substrate inhibition effects were shown with higher concentrations of all three acceptors. Manganese was shown to be necessary for galactose incorporation. A higher concentration of manganese afforded some protection from substrate inhibition by acceptors, but at the same time was itself inhibitory. All three acceptors competed with one another for galactose incorporation, indicating that a single enzyme catalyzed the transfer of galactose for all acceptors.
将肿瘤小鼠肥大细胞的悬浮培养物用于获取大量均匀的细胞群体，作为细胞分级分离的起始材料。通过对既定技术进行轻微修改制备高尔基体组分，并通过电子显微镜进行鉴定。使用卵清蛋白、去唾液酸去半乳糖化的类粘蛋白和N - 乙酰葡糖胺作为半乳糖受体来测定半乳糖基转移酶活性，结果表明高尔基体组分的比活性相对于匀浆有所富集。对高尔基体半乳糖基转移酶进行了详细研究。确定了最佳半乳糖掺入的受体浓度，并显示出所有三种受体浓度较高时均存在底物抑制作用。结果表明锰对于半乳糖掺入是必需的。较高浓度的锰可提供一定程度的保护，防止受体产生底物抑制，但同时其本身也具有抑制作用。所有三种受体相互竞争半乳糖掺入，这表明单一酶催化所有受体的半乳糖转移。
