Biosynthesis of the N-linked glycan in Campylobacter jejuni and addition onto protein through Block transfer
Biosynthesis of the N-linked glycan in Campylobacter jejuni and addition onto protein through Block transfer
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DOI：
10.1128/jb.188.7.2427-2434.2006
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发表时间：
2006-04-01
影响因子：
3.2
通讯作者：
Szymanski, CM
中科院分区：
文献类型：
--
作者：
Kelly, J;Jarrell, H;Szymanski, CM
在真核生物中，N-连接蛋白糖基化是一种普遍的修饰，包括加入预先形成的寡糖来选择ASN-Xaa-Ser/Thr基序，并影响多个生物事件。我们最近证明了空肠弯曲菌是第一个拥有N-连接的糖链途径的细菌成员。本研究应用高分辨率魔角旋转核磁共振技术(HR-MAS核磁共振)对C-空肠N-葡聚糖的体内生物合成进行了探测和定量。为了证实HR-MAS的核磁共振结果，筛选了糖基化突变体以确定鸡的定植能力，并用质谱仪和凝集素印迹法检测了糖蛋白。与真核生物的机制一致，联合数据表明细菌多糖是整体组装的，强调了蛋白质N糖基化的进化保守性。我们还发现，在所考察的条件下，PglG在糖的生物合成中不起作用，PglI是葡萄糖转移酶和可能的ABC转运蛋白，WlaB(更名为PgkK)是糖组装所必需的。这些研究为细菌中N-连接蛋白糖基化的机制奠定了基础，并为研究蛋白质糖基化和糖工程的开发提供了一个简单的模型系统。
In eukaryotes, N-linked protein glycosylation is a universal modification involving addition of preformed oligosaccharides to select Asn-Xaa-Ser/Thr motifs and influencing multiple biological events. We recently demonstrated that Campylobacter jejuni is the first member of the Bacteria to possess an N-linked glycan pathway. In this study, high-resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR) was applied to probe and quantitate C jejuni N-glycan biosynthesis in vivo. To confirm HR-MAS NMR findings, glycosylation mutants were screened for chicken colonization potential, and glycoproteins were examined by mass spectrometry and lectin blotting. Consistent with the mechanism in eukaryotes, the combined data indicate that bacterial glycans are assembled en bloc, emphasizing the evolutionary conservation of protein N glycosylation. We also show that under the conditions examined, PglG plays no role in glycan biosynthesis, PglI is the glucosyltransferase and the putative ABC transporter, and WlaB (renamed PgkK) is required for glycan assembly. These studies underpin the mechanism of N-linked protein glycosylation in Bacteria and provide a simple model system for investigating protein glycosylation and for exploitation in glycoengineering.