Enzymically inactive members of the trans-sialidase family from Trypanosoma cruzi display β-galactose binding activity
Enzymically inactive members of the trans-sialidase family from Trypanosoma cruzi display β-galactose binding activity
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DOI：
10.1093/glycob/9.6.581
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发表时间：
1999-06-01
期刊：
影响因子：
4.3
通讯作者：
Frasch, ACC
中科院分区：
文献类型：
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作者：
Cremona, ML;Campetella, O;Frasch, ACC
反式唾液酸酶是一种独特的唾液酸酶，它不是水解唾液酸，而是优先将单糖转移到糖蛋白和糖脂中的末端-半乳糖，这种酶最初在锥虫中发现，属于一个大家族的蛋白质。一些家族成员缺乏这种酶的活性。他们还没有被赋予任何职能。本文研究了反式唾液酸酶家族非活性成员的基因拷贝数和可能的功能。结果表明，编码无活性成员的基因并不少，而是与编码活性尾苷酶的基因存在相同的拷贝数（每单倍体基因组60-80个）。纯化重组失活蛋白，测定其唾液酸和半乳糖的凝集活性。酶失活的反式唾液酸酶被发现能凝集去唾液酸化的红细胞，但不凝集未经处理的红细胞。用小鼠和兔红细胞进行的实验表明，失活的反式唾液酸酶与β而不是α末端半乳糖结合，这与活性反式唾液酸酶所要求的特异性相同。一个重组分子通过单个氨基酸的突变而失去酶活性，但也保留了半乳糖的结合活性，这种结合是由乳糖竞争的，并且依赖于蛋白质天然构象的保存，因此，至少一些反式唾液酸酶家族中失去酶功能的分子仍然保留了其gal结合特性，并且可能在寄生虫-宿主相互作用中具有凝集素的功能。
trans-sialidase is a unique sialidase in that, instead of hydrolizing sialic acid, it preferentially transfers the monosaccharide to a terminal beta-galactose in glycoproteins and glycolipids, This enzyme, originally identified in Trypanosoma crud, belongs to a large family of proteins. Some members of the family lack the enzymatic activity. No function has been yet assigned to them. In this work, the gene copy number and the possible function of inactive members of the trans-sialidase family was studied. It is shown that genes encoding inactive members are not a few, but rather, are present in the same copy number (60-80 per haploid genome) as those encoding active trails-sialidases. Recombinant inactive proteins were purified and assayed for sialic acid and galactose binding activity in agglutination tests. The enzymatically inactive trans-sialidases were found to agglutinate de-sialylated erythrocytes but not untreated red blood cells. Assays made with mouse and rabbit red blood cells suggest that inactive trans-sialidases bind to beta, rather than alpha, terminal galactoses, the same specificity required by active trans-sialidases. A recombinant molecule that was made enzymatically inactive through a mutation ire a single amino acid also retained the galactose binding activity, The binding was competed by lactose and was dependent on conservation of the protein native conformation, Therefore, at least some molecules in the trans-sialidase family that have lost their enzymatic function still retain their Gal-binding properties and might have a function as lectins in the parasite-host interaction.