The structure of a neural specific carbohydrate epitope of horseradish peroxidase recognized by anti-horseradish peroxidase antiserum.
The structure of a neural specific carbohydrate epitope of horseradish peroxidase recognized by anti-horseradish peroxidase antiserum.
复制标题
抗辣根过氧化物酶抗血清识别的辣根过氧化物酶神经特异性碳水化合物表位的结构。
DOI：
--
复制
发表时间：
1991
影响因子：
4.8
通讯作者：
T. Kawasaki
中科院分区：
文献类型：
--
作者：
A. Kurosaka;A. Yano;N. Itoh;Y. Kuroda;T. Nakagawa;T. Kawasaki
辣根过氧化物酶（HRP）抗血清识别果蝇和其他昆虫的神经特异性碳水化合物抗原。将HRP糖肽与牛血清白蛋白偶联，包被于威尔斯微孔板上，再与羊抗HRP抗血清反应，用酶联免疫吸附法测定HRP糖肽与抗HRP抗血清识别的表位活性。通过杏仁糖肽酶A消化HRP胃蛋白酶底物释放的HRP糖部分进行吡啶胺化。采用反相高效液相色谱法将吡啶氨基寡糖分离成7个组分。主要部分，其中包括约80%的总糖，强烈反应与抗HRP抗血清。通过糖组成分析和600-MHz 1H NMR光谱测定该级分的碳水化合物结构如下：Man α 1-6（Man α 1-3）（Xyl β 1-2）Man β 1-4GlcNAc β 1-4（Fuc α 1-3）GlcNAc。通过外切糖苷酶消化和部分酸水解从主要部分获得的各种寡糖衍生物与抗HRP抗血清的反应性分析表明，α 1- 6-连接的甘露糖和α 1- 3-连接的岩藻糖主要参与表位结构。
Antiserum raised against horseradish peroxidase (HRP) recognizes a neural specific carbohydrate antigen in Drosophila and other insects. The epitopic activity of the carbohydrate moiety of HRP recognized by anti-HRP antiserum was measured by a newly developed enzyme-linked immunosorbent assay, in which HRP glycopeptides conjugated with bovine serum albumin were coated onto the wells and then reacted with goat anti-HRP antiserum. HRP sugar moieties released by almond glycopeptidase A digestion of HRP pepsin digests were subjected to pyridylamination. Pyridylamino oligosaccharides were separated into seven fractions by reverse-phase high performance liquid chromatography. The major fraction, which comprised about 80% of the total sugars, reacted strongly with anti-HRP antiserum. The carbohydrate structure of this fraction was determined by sugar composition analysis and 600-MHz 1H NMR spectroscopy as follows: Man alpha 1----6(Man alpha 1----3)(Xyl beta 1----2)Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----3)GlcNAc. Analyses of reactivity with anti-HRP antiserum of various oligosaccharide derivatives obtained from the major fraction by exoglycosidase digestion and partial acid hydrolysis indicated that alpha 1----6-linked mannose and alpha 1----3-linked fucose are predominantly involved in the epitopic structure.