Four N-linked glycosylation sites in human Toll-like receptor 2 cooperate to direct efficient biosynthesis and secretion
Four N-linked glycosylation sites in human Toll-like receptor 2 cooperate to direct efficient biosynthesis and secretion
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DOI：
10.1074/jbc.m403830200
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发表时间：
2004-08-13
影响因子：
4.8
通讯作者：
Gay, NJ
中科院分区：
文献类型：
--
作者：
Weber, ANR;Morse, MA;Gay, NJ
大多数高等生物都有一个先天免疫防御系统，该系统由一组进化相关的生殖系编码受体介导，即所谓的Toll样受体。在哺乳动物中，Toll样受体响应病原体相关的微生物结构发出信号。例如，Toll样受体2似乎介导对细菌肽聚糖和酰化脂蛋白的应答，Toll样受体4介导对细菌脂多糖的应答。然而，对这些结构的识别所依据的结构原理知之甚少。Toll样受体在其细胞外结构域中具有富含亮氨酸的重复序列，因此被认为采用螺线管结构，类似于在血小板糖蛋白Ib中发现的螺线管结构。此外，所有Toll样受体都含有N-连接的糖基化共有位点，Toll样受体4需要糖基化才能发挥功能。Toll样受体糖基化也可能影响受体表面表现、运输和模式识别。使用圆二色光谱，我们在这里表明，纯化的人Toll样受体2和4蛋白具有类似于糖蛋白Ib的二级结构内容。我们还分析了其他人Toll样受体的胞外结构域中共有糖基化位点的位置。我们发现，在不同的Toll样受体的网站之间的位置和程度的保守性有显着差异。使用定点突变，我们发现，在Toll样受体2胞外结构域的所有四个预测的糖基化位点被取代，虽然一个网站是低效的核心糖基化和它的去除大大影响分泌。剩余的Toll样受体2糖基化位点也有助于有效的蛋白质分泌，尽管程度较低。
Most higher organisms have a system of innate immune defense that is mediated by a group of evolutionarily related, germ line-encoded receptors, so-called Toll-like receptors. In mammals Toll-like receptors signal in response to pathogen-associated microbial structures. For example, Toll-like receptor 2 appears to mediate responses to bacterial peptidoglycan and acylated lipoproteins and Toll-like receptor 4 to bacterial lipopolysaccharide. However, the structural principles that underlie recognition of these structures are poorly understood. Toll-like receptors have leucine-rich repeats in their extracellular domains and are thus believed to adopt solenoid structures, similar to that found in platelet glycoprotein Ib. Additionally, all Toll-like receptors contain N-linked glycosylation consensus sites, and Toll-like receptor 4 requires glycosylation for function. Toll-like receptor glycosylation is also likely to influence receptor surface representation, trafficking, and pattern recognition. Using circular dichroism spectroscopy, we show here that purified human Toll-like receptor 2 and 4 proteins have secondary structure contents similar to glycoprotein Ib. We have also analyzed where consensus glycosylation sites are located in the extracellular domains of other human Toll-like receptors. We found that there are significant differences in the location and degree of conservation between sites in different Toll-like receptors. Using site-directed mutagenesis, we have found that in Toll-like receptor 2 extracellular domain all four predicted glycosylation sites are substituted, although one site is inefficiently core-glycosylated and its removal drastically affects secretion. The remaining Toll-like receptor 2 glycosylation sites also contribute to efficient protein secretion, albeit to a lesser degree.