Cellular localization and substrate specificity of isoelectric forms of human liver neuraminidase activity.
Cellular localization and substrate specificity of isoelectric forms of human liver neuraminidase activity.
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人肝神经氨酸酶活性等电形式的细胞定位和底物特异性。
DOI：
10.1042/bj2410137
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Alhadeff,JA
中科院分区：
文献类型：
--
作者：
Spaltro,J;Alhadeff,JA
人肝脏神经氨酸酶的四种主要等电形式（pI 值在 3.4 和 4.8 之间）已通过制备型等电聚焦分离出来，并使用糖蛋白、糖肽、寡糖和神经节苷脂天然底物对其底物特异性进行了表征。所有形式都表现出相当广泛的连接特异性，并且能够水解唾液酸糖苷连接的α2-3、α2-6和α2-8，尽管发现每种形式的底物水解速率不同。酸性最强的 1 型 (pI 3.4) 对唾液酸乳糖最有活性，而 2 型 (pI 3.9) 和 3 (pI 4.4) 对疏水性更强的神经节苷脂底物最有活性。 4 型 (pI 4.8) 对低 Mr 亲水底物（胎球蛋白糖肽、唾液酸乳糖）最有活性。每种形式对糖蛋白胎球蛋白的活性均低于对源自胎球蛋白的糖肽的活性。从新鲜人肝组织中制备富含细胞器的级分，并在质膜、微粒体、溶酶体和胞浆制剂中恢复对 2'-(4-甲基伞形基)-α-D-N-乙酰神经氨酸的神经氨酸酶活性。在这些制剂中恢复的神经氨酸酶活性的等电聚焦导致每种制剂显着不同的等电图（形式的数量、相对量和pI值）。等电形式的不同底物特异性和在亚细胞富集级分中恢复的神经氨酸酶活性的不同等电聚焦曲线表明，具有广泛但明确的底物特异性的特定等电形式在细胞内的不同位点富集。
The four major isoelectric forms of human liver neuraminidase (with pI values between 3.4 and 4.8) have been isolated by preparative isoelectric focusing and characterized with regard to their substrate specificity using glycoprotein, glycopeptide, oligosaccharide and ganglioside natural substrates. All forms exhibited a rather broad linkage specificity and were capable of hydrolyzing sialic acid glycosidically linked alpha 2-3, alpha 2-6 and alpha 2-8, although differential rates of hydrolysis of the substrates were found for each form. The most acidic form 1 (pI 3.4) was most active on sialyl-lactose, whereas form 2 (pI 3.9) and 3 (pI 4.4) were most active on the more hydrophobic ganglioside substrates. Form 4 (pI 4.8) was most active on the low-Mr hydrophilic substrates (fetuin glycopeptide, sialyl-lactose). Each form was less active on the glycoprotein fetuin than on a glycopeptide derived from fetuin. Organelle-enriched fractions were prepared from fresh human liver tissue and neuraminidase activity on 2′-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminic acid was recovered in plasma membrane, microsomal, lysosomal and cytosolic preparations. Isoelectric focusing of the neuraminidase activity recovered in each of these preparations resulted in significantly different isoelectric profiles (number, relative amounts and pI values of forms) for each preparation. The differential substrate specificity of the isoelectric forms and the different isoelectric focusing profiles of neuraminidase activity recovered in subcellular-enriched fractions suggest that specific isoelectric forms with broad but defined substrate specificity are enriched at separate sites within the cell.