Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

doi:10.1016/j.jia.2023.07.039

Journal of Integrative Agriculture ›› 2023, Vol. 22 ›› Issue (9): 2834-2847.DOI: 10.1016/j.jia.2023.07.039

收稿日期:2023-04-29 接受日期:2023-06-19 出版日期:2023-09-20 发布日期:2023-09-14

Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

SONG Jin-xing^{1, 2}, WANG Meng-xiang^{1, 2}, ZHANG Yi-xuan^{1, 2}, WAN Bo^{1, 2}, DU Yong-kun^{1, 2}, ZHUANG Guo-qing^{1, 2}, LI Zi-bin³, QIAO Song-lin⁶, GENG Rui¹, WU Ya-nan^{1, 2#}, ZHANG Gai-ping^{1, 2, 4, 5#}

¹ College of Veterinary Medicine, Henan Agricultural University, Zhengzhou 450046, P.R.China
² International Joint Research Center of National Animal Immunology, Zhengzhou 450046, P.R.China
³ College of Animal Science and Veterinary Medicine, Tianjin Agricultural University, Tianjin 300384, P.R.China
⁴ Longhu Laboratory, Zhengzhou 450046, P.R.China
⁵School of Advanced Agricultural Sciences, Peking University, Beijing 100871, P.R.China
⁶ Henan Provincial Key Laboratory of Animal Immunology, Henan Academy of Agricultural Sciences, Zhengzhou 450002, P.R.China

Received:2023-04-29 Accepted:2023-06-19 Online:2023-09-20 Published:2023-09-14
About author:SONG Jin-xing, Tel: +86-371-56990163, E-mail: 1010746220@qq.com; #Correspondence WU Ya-nan, E-mail: wlyananjiayou@yeah.net; ZHANG Gai-ping, E-mail: zhanggaip@126.com
Supported by:
This work was supported by the National Natural Science Foundation of China (31941001 and 32002292), the Major Science and Technology Project of Henan Province, China (221100110600) and the Natural Science Foundation of Henan Province (202300410199).

摘要/Abstract

摘要：

非洲猪瘟病毒(ASFV)是一种致命的病原体，对全球养猪业造成灾难性的社会经济影响。到目前为止，还没有获得许可的预防性疫苗。目前对非洲猪瘟病毒主要免疫原和关键抗原表位图谱的了解有限。因此，功能性单克隆抗体(mAb)和表位图谱的研究对我们理解免疫反应和设计改进的疫苗、治疗和诊断至关重要。本研究基于自然感染ASFV的康复期PBMCs构建的ASFV抗体噬菌体展示文库，筛选出一株抗ASFV主要衣壳蛋白(p72)单链抗体，并与IgG Fc片段融合(scFv-83-Fc)，可以特异性识别ASFV Pig/HLJ/2018毒株。进一步，使用scFv-83-Fc单克隆抗体为靶点，我们从M13噬菌体展示随机肽库中鉴定到p72的一段保守表位肽（²²¹MTGYKH²²⁶）。此外，流式细胞术和细胞摄取实验表明，该表位肽在体外能显著促进BMDC成熟，并能被DC细胞有效摄取，这可能意味着其在疫苗和诊断试剂开发中的潜在应用前景。总之，本研究为确定ASFV疫苗开发的靶点提供了一个有价值的平台，并有助于亚单位疫苗和诊断试剂的优化设计。

关键词:

Abstract:

African swine fever virus (ASFV) is a lethal pathogen that causes severe threats to the global swine industry and it has already had catastrophic socio-economic effects. To date, no licensed prophylactic vaccine exists. Limited knowledge exists about the major immunogens of ASFV and the epitope mapping of the key antigens. As such, there is a considerable requirement to understand the functional monoclonal antibodies (mAbs) and the epitope mapping may be of utmost importance in our understanding of immune responses and designing improved vaccines, therapeutics, and diagnostics. In this study, we generated an ASFV antibody phage-display library from ASFV convalescent swine PBMCs, further screened a specific ASFV major capsid protein (p72) single-chain antibody and fused with an IgG Fc fragment (scFv-83-Fc), which is a specific recognition antibody against ASFV Pig/HLJ/2018 strain. Using the scFv-83-Fc mAb, we selected a conserved epitope peptide (²²¹MTGYKH²²⁶) of p72 retrieved from a phage-displayed random peptide library. Moreover, flow cytometry and cell uptake experiments demonstrated that the epitope peptide can significantly promote BMDCs maturation in vitro and could be effectively uptaken by DCs, which indicated its potential application in vaccine and diagnostic reagent development. Overall, this study provided a valuable platform for identifying targets for ASFV vaccine development, as well as to facilitate the optimization design of subunit vaccine and diagnostic reagents

Key words: ASFV , phage display antibody library , single chain antibody , p72 , epitope

SONG Jin-xing, WANG Meng-xiang, ZHANG Yi-xuan, WAN Bo, DU Yong-kun, ZHUANG Guo-qing, LI Zi-bin, QIAO Song-lin, GENG Rui, WU Ya-nan, ZHANG Gai-ping. [J]. Journal of Integrative Agriculture, 2023, 22(9): 2834-2847.

SONG Jin-xing, WANG Meng-xiang, ZHANG Yi-xuan, WAN Bo, DU Yong-kun, ZHUANG Guo-qing, LI Zi-bin, QIAO Song-lin, GENG Rui, WU Ya-nan, ZHANG Gai-ping. Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library[J]. Journal of Integrative Agriculture, 2023, 22(9): 2834-2847.

参考文献

Ahamadi-Fesharaki R, Fateh A, Vaziri F, Solgi G, Siadat S, Mahboudi F, Rahimi-Jamnani F J M. 2019. Single-chain variable fragment-based bispecific antibodies: Hitting two targets with one sophisticated arrow. Molecular Therapy-Oncolytics, 14, 38–56.

Arias M, de la Torre A, Dixon L, Gallardo C, Jori F, Laddomada A, Martins C, Parkhouse R M, Revilla Y, Rodriguez F, Jose M, Sanchez V. 2017. Approaches and perspectives for development of African swine fever virus vaccines. Vaccines, 5, 35.

Belabed M, Mauvais F X, Maschalidi S, Kurowska M, Goudin N, Huang J D, Fischer A, de Saint Basile G, Endert P, Sepulveda F E, Ménasché G. 2020. Kinesin-1 regulates antigen cross-presentation through the scission of tubulations from early endosomes in dendritic cells. Nature Communications, 11, 1817.

Bradbury A R, Sidhu S, Dubel S, McCafferty J. 2011. Beyond natural antibodies: The power of in vitro display technologies. Nature Biotechnology, 29, 245–254.

Brähler S, Zinselmeyer B H, Raju S, Nitschke M, Suleiman H, Saunders B T, Johnson M W, Böhner A M C, Viehmann S F, Theisen D J, Kretzer N M, Briseño C G, Zaitsev K, Ornatsky O, Chang Q, Carrero J A, Kopp J B, Artyomov M N, Kurts C, Murphy K M, et al. 2018. Opposing roles of dendritic cell subsets in experimental GN. Journal of the American Society of Nephrology, 29, 138–154.

Chapman D A G, Tcherepanov V, Upton C, Dixon L K. 2008. Comparison of the genome sequences of non-pathogenic and pathogenic African swine fever virus isolates. Journal of General Virology, 89, 397–408.

Dixon L K, Stahl K, Jori F, Vial L, Pfeiffer D U. 2020. African swine fever epidemiology and control. Annual Review of Animal Biosciences, 8, 221–246.

Dixon L K, Sun H, Roberts H. 2019. African swine fever. Antiviral Research, 165, 34–41.

Escribano J, Galindo I, Alonso C J V. 2013. Antibody-mediated neutralization of African swine fever virus: Myths and facts. Virus Research, 173, 101–109.

Geng R, Sun Y, Li R, Yang J, Ma H, Qiao Z, Lu Q, Qiao S, Zhang G. 2022. Development of a p72 trimer-based colloidal gold strip for detection of antibodies against African swine fever virus. Applied Microbiology and Biotechnology, 106, 2703–2714.

Glockshuber R, Malia M, Pfitzinger I, Plückthun A J B. 1990. A comparison of strategies to stabilize immunoglobulin Fv-fragments. Biochemistry, 29, 1362–1367.

Gómez-Puertas P, Rodríguez F, Oviedo J, Ramiro-Ibáñez F, Ruiz-Gonzalvo F, Alonso C, Escribano J J J. 1996. Neutralizing antibodies to different proteins of African swine fever virus inhibit both virus attachment and internalization. Virology Journal, 70, 5689–5694.

Heimerman M, Murgia M, Wu P, Lowe A, Jia W, Rowland R J J. 2018. Linear epitopes in African swine fever virus p72 recognized by monoclonal antibodies prepared against baculovirus-expressed antigen. Journal of Veterinary Diagnostic Investigation, 30, 406–412.

Holliger P, Hudson P J. 2005. Engineered antibody fragments and the rise of single domains. Nature Biotechnology, 23, 1126–1136.

Iyer L M, Aravind L, Koonin E V. 2001. Common origin of four diverse families of large eukaryotic DNA viruses. Virology Journal, 75, 11720–11734.

Iyer L M, Balaji S, Koonin E V, Aravind L. 2006. Evolutionary genomics of nucleo-cytoplasmic large DNA viruses. Virus Research, 117, 156–184.

Jia N, Ou Y, Pejsak Z, Zhang Y, Zhang J. 2017. Roles of African swine fever virus structural proteins in viral infection. Journal of Veterinary Research, 61, 135–143.

Ledsgaard L, Ljungars A, Rimbault C, Sorensen C V, Tulika T, Wade J, Wouters Y, McCafferty J, Laustsen A H. 2022. Advances in antibody phage display technology. Drug Discovery Today, 27, 2151–2169.

Li K, Zettlitz K, Lipianskaya J, Zhou Y, Marks J, Mallick P, Reiter R, Wu A J P. 2015. A fully human scFv phage display library for rapid antibody fragment reformatting. Protein Engineering Design & Selection, 28, 307–316.

Liu F, Guan Z, Liu Y, Li J, Liu C, Gao Y, Ma Y, Feng J, Shen B, Yang G. 2021. Identification of a human anti-alpha-toxin monoclonal antibody against Staphylococcus aureus infection. Frontiers in Microbiology, 12, 692279.

Liu S, Luo Y, Wang Y, Li S, Zhao Z, Bi Y, Sun J, Peng R, Song H, Zhu D, Sun Y, Li S, Zhang L, Wang W, Sun Y, Qi J, Yan J, Shi Y, Zhang X, Wang P, et al. 2019. Cryo-EM structure of the African swine fever virus. Cell Host Microbe, 26, 836–843.

Neilan J G, Zsak L, Lu Z, Burrage T G, Kutish G F, Rock D L. 2004. Neutralizing antibodies to African swine fever virus proteins p30, p54, and p72 are not sufficient for antibody-mediated protection. Virology, 319, 337–342.

Onisk D V, Kutish G, Kramer E, Irusta P, Rock D L. 1994. Passively transferred African swine fever virus antibodies protect swine against lethal infection. Virology, 198, 350–354.

Pan Y, Du J, Liu J, Wu H, Gui F, Zhang N, Deng X, Song G, Li Y, Lu J, Wu X, Zhan S, Jing Z, Wang J, Yang Y, Liu J, Chen Y, Chen Q, Zhang H, Hu H, et al. 2021. Screening of potent neutralizing antibodies against SARS-CoV-2 using convalescent patients-derived phage-display libraries. Cell Discovery, 7, 57.

Revilla Y, Perez-Nunez D, Richt J A. 2018. African swine fever virus biology and vaccine approaches. Advances in Virus Research, 100, 41–74.

Sanchez E G, Perez-Nunez D, Revilla Y. 2017. Mechanisms of entry and endosomal pathway of African swine fever virus. Vaccines, 5, 42.

Sánchez-Cordón P, Montoya M, Reis A, Dixon L J V. 2018. African swine fever: A re-emerging viral disease threatening the global pig industry. Veterinary Journal, 233, 41–48.

Sheedy C, MacKenzie C, Hall J J. 2007. Isolation and affinity maturation of hapten-specific antibodies. Biotechnology Advances, 25, 333–352.

Song J, Wang M, Du Y, Wan B, Zhang A, Zhang Y, Zhuang G, Ji P, Wu Y, Zhang G. 2023. Identification of a linear B-cell epitope on the African swine fever virus CD2v protein. International Journal of Biological Macromolecules, 232, 123264.

Stefan M A, Light Y K, Schwedler J L, McIlroy P R, Courtney C M, Saada E A, Thatcher C E, Phillips A M, Bourguet F A, Mageeney C M, McCloy S A, Collette N M, Negrete O A, Schoeniger J S, Weilhammer D R, Harmon B. 2021. Development of potent and effective synthetic SARS-CoV-2 neutralizing nanobodies. mAbs, 13, 1958663.

Wang L, Luo Y, Zhao Y, Gao G, Bi Y, Qiu H J T. 2020. Comparative genomic analysis reveals an ‘open’ pan-genome of African swine fever virus. Transboundary and Emerging Diseases, 67, 1553–1562.

Wang N, Zhao D, Wang J, Zhang Y, Wang M, Gao Y, Li F, Wang J, Bu Z, Rao Z, Wang X. 2019. Architecture of African swine fever virus and implications for viral assembly. Science, 2019, 640–644.

Weisser N, Hall J J B. 2009. Applications of single-chain variable fragment antibodies in therapeutics and diagnostics. Biotechnology Advances, 27, 502–520.

Wen X, He X, Zhang X, Zhang X, Liu L, Guan Y, Zhang Y, Bu Z. 2019. Genome sequences derived from pig and dried blood pig feed samples provide important insights into the transmission of African swine fever virus in China in 2018. Emerging Microbes & Infections, 8, 303–306.

[1]	Yuan Gao, Fuxia Bai, Qi Zhang, Xiaoya An, Zhaofei Wang, Chuzhao Lei, Ruihua Dang. 基于单细胞转录组测序的公牛精子发生过程中动态转录图谱及新标记基因分析[J]. Journal of Integrative Agriculture, 2024, 23(7): 2362-2378.
[2]	. 优化耕作方式提高了机插水稻的产量并降低了稻田的温室气体排放[J]. Journal of Integrative Agriculture, 2024, 23(4): 1150-1163.
[3]	. 综合转录组学和代谢组学鉴定调控刺梨槲皮素衍生物合成的关键基因[J]. Journal of Integrative Agriculture, 2024, 23(3): 876-887.
[4]	. 妇女赋权和食物消费：来自坦桑尼亚女性户主家庭的证据[J]. Journal of Integrative Agriculture, 2024, 23(2): 457-467.
[5]	. 高温对玉米节间可塑性和株型影响的时序效应[J]. Journal of Integrative Agriculture, 2024, 23(2): 551-565.
[6]	. SUPER WOMAN 2 (SPW2) 通过抑制花同源异型基因OsMADS3、OsMADS58、OsMADS13和DROOPING LEAF的表达维持小穗的器官特征 [J]. Journal of Integrative Agriculture, 2024, 23(1): 59-76.
[7]	. 深松提高旱地小麦产量、土壤有机碳和速效养分含量[J]. Journal of Integrative Agriculture, 2024, 23(1): 251-266.
[8]	WANG Xue-feng, SHAO Dong-nan, LIANG Qian, FENG Xiao-kang, ZHU Qian-hao, YANG Yong-lin, LIU Feng, ZHANG Xin-yu, LI Yan-jun, SUN Jie, XUE Fei. 一个编码B-BOX蛋白的GhDR基因上的2bp移码缺失与陆地棉矮杆红叶性状共分离[J]. Journal of Integrative Agriculture, 2023, 22(7): 2000-2014.
[9]	GAO Yue, LUO Jian, SUN Yue, ZHANG Hua-wei, ZHANG Da-xia, LIU Feng, MU Wei, LI Bei-xing. 高效氯氟氰菊酯微囊的光敏性和粒径精准组合协同作用产生了更好的杀虫效果[J]. Journal of Integrative Agriculture, 2023, 22(5): 1477-1488.
[10]	. JIA-2022-0121 Halloween基因AhCYP307A2和AhCYP314A1调控莲草直胸跳甲(鞘翅目:叶甲科)末龄幼虫-蛹-成虫转变，卵巢发育和卵子发生[J]. Journal of Integrative Agriculture, 2023, 22(3): 812-824.
[11]	. JIA-2021-1944 水稻新型maspardin 蛋白基因OsMas1通过介导ABA信号途径调控其耐盐性和抗旱性研究[J]. Journal of Integrative Agriculture, 2023, 22(2): 341-359.
[12]	. JIA-2021-1855 真菌生物农药持续控制苜蓿害虫提升饲草品质研究[J]. Journal of Integrative Agriculture, 2023, 22(1): 185-194.
[13]	. JIA-2022-0156 PpMAPK6与PpDAM6相互作用调控桃芽内休眠解除[J]. Journal of Integrative Agriculture, 2023, 22(1): 139-148.
[14]	. 梨种质资源果实表型性状遗传多样性分析[J]. Journal of Integrative Agriculture, 2022, 21(8): 2275-2290.
[15]	. JIA-2021-0997 HBP1通过直接增强JAK2的表达来激活STAT3信号通路从而抑制鸡前脂肪细胞分化[J]. Journal of Integrative Agriculture, 2022, 21(6): 1740-1754.

Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics