GLYCOSIDASES OF EHRLICH ASCITES TUMOR-CELLS AND ASCITIC FLUID PURIFICATION AND SUBSTRATE-SPECIFICITY OF ALPHA-N-ACETYLGALACTOSAMINIDASE AND ALPHA-GALACTOSIDASE - COMPARISON WITH COFFEE BEAN ALPHA-GALACTOSIDASE
GLYCOSIDASES OF EHRLICH ASCITES TUMOR-CELLS AND ASCITIC FLUID PURIFICATION AND SUBSTRATE-SPECIFICITY OF ALPHA-N-ACETYLGALACTOSAMINIDASE AND ALPHA-GALACTOSIDASE - COMPARISON WITH COFFEE BEAN ALPHA-GALACTOSIDASE
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DOI：
10.1016/0003-9861(90)90518-4
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发表时间：
1990-07-01
影响因子：
3.9
通讯作者：
GOLDSTEIN, IJ
中科院分区：
文献类型：
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作者：
YAGI, F;ECKHARDT, AE;GOLDSTEIN, IJ
测定埃利希腹水肿瘤细胞和腹水的糖苷酶活性。α-半乳糖苷酶，和β-半乳糖苷酶。半乳糖苷酶，α-和β-甘露糖苷酶，α- N-乙酰氨基半乳糖苷酶，和β- N-乙酰氨基葡萄糖苷酶活性检测使用对硝基苯基糖苷作为底物。α-半乳糖苷酶和α- N-乙酰氨基半乳糖苷酶是在ε-H2O上从埃利希腹水肿瘤细胞中分离的。氨基己酰半乳糖胺-琼脂糖凝胶。α-半乳糖苷酶被纯化160，000倍，并且不含其他糖苷酶活性。α- N-乙酰氨基半乳糖苷酶也纯化了160，000倍，但表现出弱的α-半乳糖苷酶活性。半乳糖苷酶活性似乎是这种酶固有的。α-半乳糖苷酶的底物特异性用12种底物研究了半乳糖苷酶，并与相应的咖啡豆酶进行了比较。埃利希细胞α-β的pH最适值半乳糖苷酶的pH值集中在4.5附近，与底物无关，而咖啡豆酶对PNP-α-半乳糖苷酶的最适pH值为4.5。Gal为6.0，比咖啡豆酶的其他底物高1.5个pH单位。发现α的情况相反- N-乙酰氨基半乳糖苷酶：PNP-α-水解的最适pH GalNAc为3.6，低于GalNAc α 1，3Gal水解所需的pH 4.5。咖啡豆α半乳糖苷酶显示出相对宽的底物特异性，这表明它适合于切割许多种类的末端α-半乳糖苷酶。半乳糖基连接。另一方面，埃利希α-半乳糖苷酶似乎是相当狭窄的。该酶对末端α-氨基甲酸酯具有高活性。埃利希糖蛋白和层粘连蛋白的半乳糖基键，两者都具有Gal α 1，3Gal β 1，4GlcNAc β-三糖序列。α-发现N-乙酰氨基半乳糖苷酶对血型A型二糖和三糖以及具有A型活性糖链的糖蛋白具有活性。
Ehrlich ascites tumor cells and ascitic fluid were assayed for glycosidase activity. .alpha.-Galactosidase, and .beta.-galactosidase, .alpha.- and .beta.-mannosidase, .alpha.-N-acetylgalactosaminidase, and .beta.-N-acetylglucosaminidase activities were detected using p-nitrophenyl glycosides as substrates. .alpha.-Galactosidase and .alpha.-N-acetylgalactosaminidase were isolated from Ehrlich ascites tumor cells on .epsilon.-aminocaproylgalactosylamine-Sepharose. .alpha.-Galactosidase was purified 160,000-fold and was free of other glycosidase activities. .alpha.-N-Acetylgalactosaminidase was also purified 160,000-fold but exhibited a weak .alpha.-galactosidase activity which appears to be inherent in this enzyme. Substrate specificity of the .alpha.-galactosidase was investigated with 12 substrates and compared with that of the corresponding coffee bean enzyme. The pH optimum of the Ehrlich cell .alpha.-galactosidase centered near 4.5, irrespective of substrate, whereas the pH optimum of the coffee bean enzyme for PNP-.alpha.-Gal was 6.0, which is 1.5 pH units higher than that for other substrates of the coffee bean enzyme. The reverse was found for .alpha.-N-acetylgalactosaminidase: the pH optimum for the hydrolysis of PNP-.alpha.-GalNAc was 3.6, lower than the pH 4.5 required for the hydrolysis of GalNAc.alpha.1,3Gal. Coffee bean .alpha.-galactosidase showed a relatively broad substrate specificity, suggesting that it is suited for cleaving many kinds of terminal .alpha.-galactosyl linkages. On the other hand, the substrate specificity of Ehrlich .alpha.-galactosidase appears to be quite narrow. This enzyme was highly active toward the terminal .alpha.-galactosyl linkages of Ehrlich glycoproteins and laminin, both of which possess Gal.alpha.1,3Gal.beta.1,4GlcNAc.beta.-trisaccharide sequences. The .alpha.-N-acetylgalactosaminidase was found to be active toward the blood group type A disaccharide, and trisaccharide, and glycoproteins with type A-active carbohydrate chains.