AbstractThe influenza neuraminidase (NA) is a potential target for the development of a next-generation influenza vaccine, but its antigenicity is not well understood. Here, we isolate an anti-N6 human monoclonal antibody, named 18_14D, from an H5N6 avian influenza virus (AIV) infected patient. The antibody weakly inhibits enzymatic activity but confers protection in female mice, mainly via ADCC function.

摘要流感神经氨酸酶（NA）是开发下一代流感疫苗的潜在靶标，但其抗原性尚不清楚。在这里，我们从H5N6禽流感病毒（AIV）感染的患者中分离出一种名为18'U 14D的抗N6人单克隆抗体。该抗体弱抑制酶活性，但主要通过ADCC功能赋予雌性小鼠保护作用。

The cryo-EM structure shows that 18_14D binds to a unique epitope on the lateral surface of the N6 tetramer, preventing the formation of tightly closed NA tetramers. These findings contribute to the molecular understanding of protective immune responses to NA of AIVs in humans and open an avenue for the rational design of NA-based vaccines..

cryo-EM结构表明，18'U 14D与N6四聚体侧面的独特表位结合，阻止了紧密闭合的NA四聚体的形成。这些发现有助于分子理解人类对禽流感病毒NA的保护性免疫反应，并为基于NA的疫苗的合理设计开辟了一条途径。。

IntroductionInfluenza viruses are continuing to evolve and cause seasonal flu and occasional worldwide pandemics, which have posed a great challenge to public health1,2,3,4. Influenza viruses have two major surface glycoproteins, hemagglutinin (HA), and neuraminidase (NA). HA mediates the binding of viruses to the cellular sialic acid receptors and fusion of the virion membrane with the intracellular endosome membranes5.

引言流感病毒正在不断进化，并导致季节性流感和偶尔的全球大流行，这对公共卫生构成了巨大挑战1,2,3,4。流感病毒有两种主要的表面糖蛋白，血凝素（HA）和神经氨酸酶（NA）。HA介导病毒与细胞唾液酸受体的结合以及病毒粒子膜与细胞内内体膜的融合5。

NA removes decoy receptors from mucins and cleaves off the sialic acid, allowing the release of progeny viruses from the infected cells6,7.Human protective antibodies elicited by vaccination or infection can bind to both the HA and NA proteins8,9. In the past years, the NA has been historically understudied compared to the surface protein counterpart HA.

NA从粘蛋白中去除诱饵受体并切割唾液酸，允许从感染细胞中释放后代病毒6,7。通过疫苗接种或感染引发的人类保护性抗体可与HA和NA蛋白结合8,9。在过去的几年中，与表面蛋白对应物HA相比，NA在历史上一直未被充分研究。

However, increasing studies highlighted the importance of NA-targeting antibodies and their implications for therapy and design of universal flu vaccines, as they have shown that NA-targeted immunity can confer protective efficacy against influenza virus challenge in animal models10,11,12,13. NA-targeting monoclonal antibodies (mAbs) are shown to reduce the viral loads and symptoms in infected mice and even correlate with the protection in humans14,15,16, and antigenic drifts also occurred on NA protein16,17,18,19,20.Most of the NA-targeting antibodies directly inhibit enzymatic activity of the NA via binding to enzyme active site21,22,23,24,25, or indirectly inhibit enzymatic activity of the NA through steric hindrance24,26,27,28.

然而，越来越多的研究强调了NA靶向抗体的重要性及其对通用流感疫苗治疗和设计的影响，因为他们已经证明NA靶向免疫可以在动物模型中赋予针对流感病毒攻击的保护功效10,11,12,13。NA靶向单克隆抗体（mAb）可降低感染小鼠的病毒载量和症状，甚至与人类的保护作用相关14,15,16，NA蛋白16,17,18,19,20也发生抗原漂移。大多数NA靶向抗体通过与酶活性位点21,22,23,24,25结合直接抑制NA的酶活性，或通过空间位阻间接抑制NA的酶活性24,26,27,28。

Otherwise, there are some other NA antibodies with no inhibitory activity to the NA, could also contribute to protection through antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP)16.In addition to seasonal H1N1, H3N2 influenza A.

否则，还有一些其他NA抗体对NA没有抑制活性，也可能通过抗体依赖性细胞毒性（ADCC）或抗体依赖性细胞吞噬作用（ADCP）16有助于保护。除了季节性H1N1，H3N2甲型流感。

Data availability

数据可用性

The atomic coordinates data generated in this study have been deposited in the Protein Data Bank (PDB) database under accession code 8Z4E. The corresponding electron microscopy maps have been deposited in the Electron Microscopy Data Bank (EMDB) with the accession codes EMD-39762. Other structures for analysis, including 2CML, 4QN6, 6Q23, 6PZY, 8EZ3, 8EZ7, 8E6J, 8GAV, 4QNP, 6PZW, 6U02, were obtained from the PDB.

。相应的电子显微镜图已保存在电子显微镜数据库（EMDB）中，登录号为EMD-39762。从PDB获得了其他用于分析的结构，包括2CML，4QN6，6Q23，6PZY，8EZ3，8EZ7，8E6J，8GAV，4QNP，6PZW，6U02。

Source data are provided as a Source Data file. Source data are provided with this paper..

源数据作为源数据文件提供。本文提供了源数据。。

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Download referencesAcknowledgementsWe thank all the staff members at the Center for Biological Imaging (CBI), Institute of Biophysics (IBP), Chinese Academy of Sciences (CAS) for assistance with data collection. We would like to thank Wenjuan Zhang for her help on negatively stained sampling and imaging at the Cryo-electron Microscopy Platform, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences (IGDB).

下载参考文献致谢我们感谢中国科学院生物物理研究所生物成像中心（CBI）的所有工作人员为数据收集提供的帮助。我们要感谢张文娟在中国科学院遗传与发育生物学研究所（IGDB）低温电子显微镜平台上对负染色采样和成像的帮助。

We also thank Zheng Fan, Qian Wang from the Institute of Microbiology, Chinese Academy of Sciences for help in BLI and ELLA assay. This study was supported by National Natural Science Foundation of China (NSFC) (32192452 to Y.S.), the National Key Research and Development Program of China (2023YFC2307800 to M.W.), National Natural Science Foundation of China (NSFC) (81902058 to C.S., and 32100119 to Q.P.), Shenzhen High‐level Hospital Construction Fund (23250G1001 to Y.Y.) and a grant from Sanofi Pasteur company.Author informationAuthor notesThese authors contributed equally: Min Wang, Yuan Gao, Chenguang Shen.Authors and AffiliationsCAS Key Laboratory of Pathogen Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences (CAS), Beijing, ChinaMin Wang, Yuan Gao, Qi Peng, Jinlong Cheng, George Fu Gao & Yi ShiShenzhen Key Laboratory of Pathogen and Immunity, Shenzhen Third People’s Hospital, Shenzhen, ChinaMin Wang, Chenguang Shen, Yang Yang & Yi ShiFaculty of Health Sciences, University of Macau, Macau, ChinaYuan Gao & Han-Ming ShenBSL-3 Laboratory (Guangdong), Guangdong Provincial Key Laboratory of Tropical Disease Research, School of Public Health; Southern Medical University, Guangzhou, ChinaChenguang ShenMedical School, University of Chinese Academy of Sciences, Beijing, ChinaWei Yang, George Fu Gao & Y.

我们还要感谢中国科学院微生物研究所的郑凡，钱旺在BLI和ELLA检测方面的帮助。这项研究得到了国家自然科学基金（NSFC）（32192452至Y.S.），国家重点研究发展计划（2023YFC2307800至M.W.），国家自然科学基金（NSFC）（81902058至C.S.，32100119至Q.P.），深圳市高水平医院建设基金（23250G1001至Y.Y.）和赛诺菲巴斯德公司的资助。作者信息作者注意到这些作者做出了同样的贡献：王敏，袁高，沈晨光。作者和所属单位中国科学院微生物研究所病原体微生物学和免疫学重点实验室，北京，王华民，袁高，齐鹏，程金龙，高志福和易世深圳深圳市第三人民医院病原体和免疫重点实验室，深圳，王华民，沈晨光，杨洋和易世深圳，澳门大学，澳门大学，中国元高和韩明ShenBSL-3实验室（广东），广东省热带病研究重点实验室，公共卫生学院；。

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PubMed Google ScholarContributionsY.S., G.F.G. and Y.Y. conceived the study. M.W., Y.G., C.S., W.Y., Q.P., and J.C performed the experiments. Y.S., M.W., C.S., and Y.Y. conducted the analysis. M.W., Y.G., C.S., H.S., Y.Y., and Y.S. wrote the manuscript. All authors participated in the discussion and manuscript editing.

PubMed谷歌学术贡献。S、 ，G.F.G.和Y.Y.构思了这项研究。M、 W.，Y.G.，C.S.，W.Y.，Q.P。和J.C进行了实验。Y、 S.，M.W.，C.S。和Y.Y.进行了分析。M、 W.，Y.G.，C.S.，H.S.，Y.Y。和Y.S.撰写了手稿。所有作者都参加了讨论和稿件编辑。

Y.S. supervised all the work.Corresponding authorsCorrespondence to.

Y、 美国监督了所有的工作。通讯作者通讯。

Yang Yang, George Fu Gao or Yi Shi.Ethics declarations

杨扬，乔治·富高或伊始。道德宣言

Competing interests

相互竞争的利益

A patent describing potential treatments for influenza using 18_14D was in application, and the authors listed on the application are Y.S., G.F.G., M.W., Y.G., Q.P., Y.Y., C.S. The remaining authors declare no competing interests.

一项描述使用18'U 14D治疗流感的潜在治疗方法的专利正在申请中，申请中列出的作者是Y.S.，G.F.G.，M.W.，Y.G.，Q.P.，Y.Y.，C.S。其余作者声明没有利益冲突。

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Reprints and permissionsAbout this articleCite this articleWang, M., Gao, Y., Shen, C. et al. A human monoclonal antibody targeting the monomeric N6 neuraminidase confers protection against avian H5N6 influenza virus infection.

转载和许可本文引用本文Wang，M.，Gao，Y.，Shen，C。等人。靶向单体N6神经氨酸酶的人单克隆抗体赋予针对禽H5N6流感病毒感染的保护作用。

Nat Commun 15, 8871 (2024). https://doi.org/10.1038/s41467-024-53301-6Download citationReceived: 17 May 2024Accepted: 09 October 2024Published: 15 October 2024DOI: https://doi.org/10.1038/s41467-024-53301-6Share this articleAnyone you share the following link with will be able to read this content:Get shareable linkSorry, a shareable link is not currently available for this article.Copy to clipboard.

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