Human N-acetylglucosaminyltransferase II substrate recognition uses a modular architecture that includes a convergent exosite
Human N-acetylglucosaminyltransferase II substrate recognition uses a modular architecture that includes a convergent exosite
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DOI：
10.1073/pnas.1716988115
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发表时间：
2018-05-01
影响因子：
11.1
通讯作者：
Moremen, Kelley W.
中科院分区：
文献类型：
--
作者：
Kadirvelraj, Renuka;Yang, Jeong-Yeh;Moremen, Kelley W.
Asn连接的寡糖在通过分泌途径转运期间被广泛修饰，首先通过修剪新生聚糖链，随后通过引发和延伸来自三甘露糖基聚糖核心的多个寡糖分支。修剪和分支途径的步骤是高度有序的和分层的基础上，精确的底物特异性的个别生物合成酶。复合型聚糖合成中的关键关键步骤由N-乙酰葡糖胺基转移酶II（MGAT 2）催化，MGAT 2是一种使用UDP-GlcNAc作为糖供体从三甘露糖基聚糖核心生成第二个GlcNAc β 1，2-分支的酶。我们确定了作为Mn 2 +-UDP供体类似物复合物和作为GlcNAc-Man（3）GlcNAc（2）-Asn受体复合物的人MGAT 2的结构，以揭示底物识别和催化的结构基础。该酶表现出GTA Rossmann样折叠，其采用保守的二价阳离子依赖性底物与UDP-GlcNAc供体的相互作用。MGAT 2与延伸聚糖受体的相互作用不同于其他相关的糖基转移酶。这些相互作用由结合Man-α 1，6-单糖受体的催化亚位点和结合GlcNAc-β 1，2 Man-α 1，3 Man β-底物“识别臂的远端外部位点口袋组成。“识别臂相互作用类似于高尔基体α-甘露糖苷酶II的酶-底物相互作用，这是一种糖苷水解酶，在Asn连接的聚糖生物合成途径中作用于MGAT 2之前。这些数据表明，MGAT 2的底物结合采用保守和收敛催化亚位点模块来提供底物选择性和催化。更广泛地说，MGAT 2活性位点结构展示了糖基转移酶如何产生互补的模块化模板，用于哺乳动物细胞中聚糖结构的区域特异性延伸。
Asn-linked oligosaccharides are extensively modified during transit through the secretory pathway, first by trimming of the nascent glycan chains and subsequently by initiating and extending multiple oligosaccharide branches from the trimannosyl glycan core. Trimming and branching pathway steps are highly ordered and hierarchal based on the precise substrate specificities of the individual biosynthetic enzymes. A key committed step in the synthesis of complex-type glycans is catalyzed by N-acetylglucosaminyltransferase II (MGAT2), an enzyme that generates the second GlcNAc beta 1,2-branch from the trimannosyl glycan core using UDP-GlcNAc as the sugar donor. We determined the structure of human MGAT2 as a Mn2+-UDP donor analog complex and as a GlcNAc-Man(3)GlcNAc(2)-Asn acceptor complex to reveal the structural basis for substrate recognition and catalysis. The enzyme exhibits a GTA Rossmann-like fold that employs conserved divalent cationdependent substrate interactions with the UDP-GlcNAc donor. MGAT2 interactions with the extended glycan acceptor are distinct from other related glycosyltransferases. These interactions are composed of a catalytic subsite that binds the Man-alpha 1,6-mono-saccharide acceptor and a distal exosite pocket that binds the GlcNAc-beta 1,2Man-alpha 1,3Man beta-substrate "recognition arm." Recognition arm interactions are similar to the enzyme-substrate interactions for Golgi a-mannosidase II, a glycoside hydrolase that acts just before MGAT2 in the Asn-linked glycan biosynthetic pathway. These data suggest that substrate binding by MGAT2 employs both conserved and convergent catalytic subsite modules to provide substrate selectivity and catalysis. More broadly, the MGAT2 active-site architecture demonstrates how glycosyltransferases create complementary modular templates for regiospecific extension of glycan structures in mammalian cells.