Comparative Analysis of the Glycosylation Profiles of Membrane-Anchored HIV-1 Envelope Glycoprotein Trimers and Soluble gp140
Comparative Analysis of the Glycosylation Profiles of Membrane-Anchored HIV-1 Envelope Glycoprotein Trimers and Soluble gp140
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DOI：
10.1128/jvi.00628-15
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发表时间：
2015-08-01
影响因子：
5.4
通讯作者：
Desaire, Heather
中科院分区：
文献类型：
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作者：
Go, Eden P.;Herschhorn, Alon;Desaire, Heather
人类免疫缺陷病毒1型（HIV-1）包膜糖蛋白（Env）三聚体由gp 120和gp 41亚基组成，是疫苗开发的多种策略的焦点。广泛的Env糖基化为HIV-1提供了免疫系统保护，但聚糖也是许多广泛中和抗体的结合表位的必要组分。最近的研究表明，当Env从病毒体中分离时，其糖基化谱与主要用于疫苗发现研究的可溶形式的Env（gp 120或gp 140）的糖基化谱显著不同。在这里，我们表明，外源性膜锚定的Envs，它可以在哺乳动物细胞中大量生产，也显示病毒体样聚糖的配置文件，其中糖蛋白被广泛装饰高甘露糖聚糖。此外，由于我们用高保真分析方法（糖肽分析）表征了糖基化，因此呈现了关于膜Env糖基化及其异质性的前所未有的分子细节水平。每个糖基化位点单独表征，每个Env蛋白表征约500种糖型。虽然许多位点仅含有高甘露糖聚糖，但其他位点保留复杂聚糖，导致目前无法在可溶性gp 120或gp 140制剂上模拟的聚糖谱。这些位点水平的研究对于理解天然Env三聚体上的抗体-聚糖相互作用是重要的。此外，我们报告了一个新观察到的O-连接的糖基化位点，T606，我们表明，完整的O-连接的糖基化概况的膜相关的Env是类似的可溶性gp 140。这些发现为Env糖基化提供了新的见解，并阐明了膜锚定Env和可溶性gp 140之间的关键分子水平差异。重要提示：预防人类免疫缺陷病毒1型（HIV-1）感染的疫苗应能引发与病毒膜表面包膜糖蛋白结合的抗体。包膜糖蛋白具有广泛的碳水化合物（聚糖）外壳，其中一些被病毒中和抗体识别，其中一些保护病毒免受中和抗体的侵害。我们发现HIV-1膜包膜糖蛋白具有独特的碳水化合物模式，具有许多高甘露糖聚糖，并且在某些地方具有复杂的聚糖。这种模式是非常不同的碳水化合物的档案看到一个更容易产生的可溶性版本的包膜糖蛋白。我们的研究结果提供了一个详细的表征的聚糖的天然膜包膜糖蛋白的HIV-1，碳水化合物的档案，这将是理想的模拟疫苗。
The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer, which consists of the gp120 and gp41 subunits, is the focus of multiple strategies for vaccine development. Extensive Env glycosylation provides HIV-1 with protection from the immune system, yet the glycans are also essential components of binding epitopes for numerous broadly neutralizing antibodies. Recent studies have shown that when Env is isolated from virions, its glycosylation profile differs significantly from that of soluble forms of Env (gp120 or gp140) predominantly used in vaccine discovery research. Here we show that exogenous membrane-anchored Envs, which can be produced in large quantities in mammalian cells, also display a virion-like glycan profile, where the glycoprotein is extensively decorated with high-mannose glycans. Additionally, because we characterized the glycosylation with a high-fidelity profiling method, glycopeptide analysis, an unprecedented level of molecular detail regarding membrane Env glycosylation and its heterogeneity is presented. Each glycosylation site was characterized individually, with about 500 glycoforms characterized per Env protein. While many of the sites contain exclusively high-mannose glycans, others retain complex glycans, resulting in a glycan profile that cannot currently be mimicked on soluble gp120 or gp140 preparations. These site-level studies are important for understanding antibody-glycan interactions on native Env trimers. Additionally, we report a newly observed O-linked glycosylation site, T606, and we show that the full O-linked glycosylation profile of membrane-associated Env is similar to that of soluble gp140. These findings provide new insight into Env glycosylation and clarify key molecular-level differences between membrane-anchored Env and soluble gp140.IMPORTANCEA vaccine that protects against human immunodeficiency virus type 1 (HIV-1) infection should elicit antibodies that bind to the surface envelope glycoproteins on the membrane of the virus. The envelope glycoproteins have an extensive coat of carbohydrates (glycans), some of which are recognized by virus-neutralizing antibodies and some of which protect the virus from neutralizing antibodies. We found that the HIV-1 membrane envelope glycoproteins have a unique pattern of carbohydrates, with many high-mannose glycans and also, in some places, complex glycans. This pattern was very different from the carbohydrate profile seen for a more easily produced soluble version of the envelope glycoprotein. Our results provide a detailed characterization of the glycans on the natural membrane envelope glycoproteins of HIV-1, a carbohydrate profile that would be desirable to mimic with a vaccine.