Narasimhan S
J Biol Chem. 1982 Sep 10;257(17):10235-42.
Hen oviduct membranes are shown to catalyze the following enzyme reaction: GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc-Asn + UDP-GlcNAc leads to GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-2Man alpha 1-3)GlcNAc beta 1-4)Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc-Asn + UDP. The enzyme catalyzing this reaction has been named UDP-GlcNAc:glycopeptide beta 4-N-acetylglucosaminyltransferase III (GlcNAc-transferase III) to distinguish it from two other GlcNAc-transferases (I and II) present in hen oviduct and previously described in several mammalian tissues. GlcNAc-transferases I and II, respectively, attach GlcNAc in beta 1-2 linkage to the Man alpha 1-3 and Man alpha 1-6 arms of Asn-linked oligosaccharide cores. A specific assay for GlcNAc-transferase III was devised by using concanavalin A/Sepharose columns to separate the product of transferase III from other interfering radioactive glycopeptides formed in the reaction. The specific activity of GlcNAc-transferase III in hen oviduct membranes is about 5 nmol/mg of protein/h. Substrate specificity studies have shown that GlcNAc-transferase III requires both terminal beta 1-2-linked GlcNAc residues in its substrate for maximal activity. Removal of the GlcNAc residue on the Man alpha 1-6 arm reduces activity by at least 85% and removal of both GlcNAc residues reduces activity by at least 93%. Two large scale preparations of product were subjected to high resolution proton NMR spectroscopy to establish the incorporation by the enzyme of a GlcNAc in beta 1-4 linkage to the beta-linked Man. This GlcNAc residue is called a "bisecting" GlcNAc and appears to play important control functions in the synthesis of complex N-glycosyl oligosaccharides. Several enzymes in the biosynthetic scheme are unable to act on glycopeptide substrates containing a bisecting GlcNAc residue.
GlcNAcβ1-2Manα1-6(GlcNAcβ1-2Manα1-3)Manβ1-4GlcNAcβ1-4(Fucα1-6)GlcNAc-Asn + UDP-GlcNAc 生成 GlcNAcβ1-2Manα1-6(GlcNAcβ1-2Manα1-3)GlcNAcβ1-4)Manβ1-4GlcNAcβ1-4(Fucα1-6)GlcNAc-Asn + UDP。催化此反应的酶被命名为 UDP-GlcNAc：糖肽β4-N-乙酰葡糖胺基转移酶 III（GlcNAc-转移酶 III），以将其与母鸡输卵管中存在且先前在几种哺乳动物组织中描述过的另外两种 GlcNAc-转移酶（I 和 II）区分开来。GlcNAc-转移酶 I 和 II 分别以β1-2 连接的方式将 GlcNAc 连接到天冬酰胺连接的寡糖核心的 Manα1-3 和 Manα1-6 臂上。通过使用伴刀豆球蛋白 A/琼脂糖柱将转移酶 III 的产物与反应中形成的其他干扰性放射性糖肽分离，设计了一种针对 GlcNAc-转移酶 III 的特异性测定方法。母鸡输卵管膜中 GlcNAc-转移酶 III 的比活性约为 5 nmol/mg 蛋白质/小时。底物特异性研究表明，GlcNAc-转移酶 III 在其底物中需要两个末端β1-2 连接的 GlcNAc 残基才能达到最大活性。去除 Manα1-6 臂上的 GlcNAc 残基会使活性降低至少 85%，去除两个 GlcNAc 残基会使活性降低至少 93%。对两种大规模制备的产物进行了高分辨率质子核磁共振光谱分析，以确定该酶将一个 GlcNAc 以β1-4 连接的方式连接到β连接的 Man 上。这个 GlcNAc 残基被称为“平分”GlcNAc，似乎在复杂 N-糖基寡糖的合成中起重要的控制作用。生物合成途径中的几种酶无法作用于含有平分 GlcNAc 残基的糖肽底物。
