Glycosylation Analysis of Engineered H3N2 Influenza A Virus Hemagglutinins with Sequentially Added Historically Relevant Glycosylation Sites
Glycosylation Analysis of Engineered H3N2 Influenza A Virus Hemagglutinins with Sequentially Added Historically Relevant Glycosylation Sites
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DOI：
10.1021/acs.jproteome.5b00416
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发表时间：
2015-09-01
影响因子：
4.4
通讯作者：
Cipollo, John F.
中科院分区：
文献类型：
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作者：
An, Yanming;McCullers, Jonathan A.;Cipollo, John F.
流感病毒表面糖蛋白血凝素（HA）是宿主中和抗体的主要目标。 HA的寡糖有助于HA的抗原特性。在从人畜共患宿主传染给人类后，流感病毒可以随着时间的推移获得 N-糖基化序列，作为其适应性策略的一部分。这种糖基化扩展尚未在结构水平上进行研究。在这里，我们检查了 H3N2 病毒株的 HA N-糖基化，我们对这些病毒株进行了改造，以密切模仿 1968 年至 2002 年期间从大流行 A/香港/1/68（H3N2：HK68）开始获得的糖基化位点。研究的 HA 包括 HK68 和添加了 1、2 和 4 个位点的工程形式。我们使用了：nano-LC-MSE 进行糖肽组成、序列和位点占用分析，并使用 MALDI-TOF MS 全甲基化分析来表征释放的聚糖。我们的研究表明，1) 大多数 N-序列片段的占据率 >90%，2) 聚糖的类别和复杂性因蛋白质区域的不同而不同，3) Asn 165 和 Asn 246 与 HA 和 SP-D 肺集合素之间的相互作用有关，完全是高甘露糖类型。根据这项研究和之前的报告，我们提供了关于免疫系统反应如何根据 HA 糖基化而有所不同的结构见解。
The influenza virus surface glycoprotein hemagglutinin (HA) is the major target of host neutralizing antibodies. The oligosaccharides of HA can contribute to HA's antigenic characteristics. After a leap to humans from a zoonotic host, influenza can gain N-glycosylation sequons over time as part of its fitness strategy. This glycosylation expansion has not been studied at the structural level. Here we examine HA N-glycosylation of H3N2 virus strains that we have engineered to closely mimic glycosylation sites gained between 1968 through 2002 starting with pandemic A/Hong Kong/1/68 (H3N2: HK68). HAs studied include HK68 and engineered forms with 1, 2, and 4 added sites. We have used: nano-LC-MSE for glycopeptide composition, sequence and site occupancy analysis, and MALDI-TOF MS permethylation profiling for characterization of released glycans. Our study reveals that 1) the majority of N-sequons are occupied at >90%, 2) the class and complexity of the glycans varies by region over the landscape of the proteins, 3) Asn 165 and Asn 246, which are associated with interactions between HA and SP-D lung collectin, are exclusively high mannose type. Based on this study and previous reports we provide structural insight as to how the immune system responses may differ depending on HA glycosylation.