可溶性流感病毒B/Victoria系血凝素重组蛋白的筛选

doi:10.16352/j.issn.1001-6325.2026.01.0092

基础医学与临床 ›› 2026, Vol. 46 ›› Issue (1): 92-96.doi: 10.16352/j.issn.1001-6325.2026.01.0092

可溶性流感病毒B/Victoria系血凝素重组蛋白的筛选

王景凡, 杨姣姣, 张婷, 王志荣, 许雪梅^*

中国医学科学院基础医学研究所北京协和医学院基础学院生物物理及结构生物学系,北京 100005

收稿日期:2025-07-14 修回日期:2025-10-30 出版日期:2026-01-05 发布日期:2025-12-29
通讯作者: ^*xuemeixu_pumc@163.com
基金资助:
中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-043)

Screening of soluble hemagglutinin recombinant proteins for influenza virus B/Victoria

WANG Jingfan, YANG Jiaojiao, ZHANG Ting, WANG Zhirong, XU Xuemei^*

Department of Biophysics and Structure Biology, Institute of Basic Medical Sciences CAMS, School of Basic Medicine PUMC, Beijing 100005, China

Received:2025-07-14 Revised:2025-10-30 Online:2026-01-05 Published:2025-12-29
Contact: ^*xuemeixu_pumc@163.com

摘要/Abstract

摘要： 目的筛选可溶性表达且具有血凝活性的流感毒株的B/Victoria系血凝素(HA)重组蛋白。方法以B/Darwin/7/2019(B/Victoria)毒株为基础,在HA胞外区C端分别融合亮氨酸拉链GCN4pII和GCN4pLL三聚化基序(T1和T2)、T4噬菌体Foldon的三聚化基序(T3)。获得HA突变体基因BV-T1、BV-T2和BV-T3,同时构建HA胞外区突变体基因BV-ecto作对照组,插入pFastBac1载体,利用杆状病毒-Sf9细胞表达体系,表达HA突变体蛋白,收获感染细胞培养上清。采用Strep-Tactin亲和层析纯化,对HA突变体纯化蛋白的寡聚化和血凝活性进行鉴定。结果 4种HA突变体蛋白均在上清中获得可溶性表达;BV-T2和BV-T3的三聚化程度很高,BV-T1以三聚化为主,存在少量的低聚化, BV-ecto以单体存在;BV-T1和BV-T2有血凝活性,而BV-T3和BV-ecto无血凝活性。结论可溶性表达的BV-T2 HA突变体蛋白三聚化程度好,且具有血凝活性。亮氨酸拉链三聚化基序GCN4pLL可用于B/Victoria系HA重组蛋白的可溶性表达,为可溶性B/Victoria系HA重组蛋白疫苗的研发提供参考。

关键词: 流感病毒, B/Victoria, 血凝素(HA), 可溶性表达

Abstract: Objective To screen the soluble hemagglutinin (HA) recombinant protein derived from B/Victoria lineages of influenza virus that possess hemagglutination activity. Methods Based on the B/Darwin/7/2019 (B/Victoria) strain, HA mutant genes BV-T1, BV-T2, and BV-T3 were obtained by fusing the leucine zipper GCN4pII and GCN4pLL trimerization motifs(T1 and T2), and bacteriophage T4 Foldon trimerization motifs(T3) to the C-terminus of the HA ectodomain. HA ectodomain mutant gene BV-ecto was also constructed as a control. These genes were inserted into pFastBac1 vector. The proteins were expressed using baculovirus-Sf9 insect cell expression system and culture supernatants of infected cells were harvested. Recombinant proteins were purified by Strep-Tactin affinity chromatography and subsequently characterized for their oligomerization degree and hemagglutination activity. Results All four mutant proteins were solubly expressed in the culture supernatant. BV-T2 and BV-T3 exhibited a high trimerization form, while BV-T1 formed predominantly trimers with minor low-order oligomers. BV-ecto existed as a monomer.BV-T1 and BV-T2 exhibited hemagglutination activity, in contrast, BV-T3 and BV-ecto lacked hemagglutination activity. Conclusions The soluble BV-T2 mutant protein present efficient trimer- ization and possesses hemagglutination activity.The leucine zipper trimerization motif GCN4pLL is suitable for the soluble expression of B/Victoria lineage HA recombinant protein and thus provides a reference for the development of soluble recombinant protein vaccines against B/Victoria virus.

Key words: influenza virus, B/Victoria, hemagglutinin (HA), soluble expression

中图分类号:

R373.1

王景凡, 杨姣姣, 张婷, 王志荣, 许雪梅. 可溶性流感病毒B/Victoria系血凝素重组蛋白的筛选[J]. 基础医学与临床, 2026, 46(1): 92-96.

WANG Jingfan, YANG Jiaojiao, ZHANG Ting, WANG Zhirong, XU Xuemei. Screening of soluble hemagglutinin recombinant proteins for influenza virus B/Victoria[J]. Basic & Clinical Medicine, 2026, 46(1): 92-96.

参考文献

[1]Iuliano AD, Roguski KM, Chang HH, et al. Estimates of global seasonal influenza-associated respiratory mortality: a modelling study[J]. Lancet, 2018, 391:1285-1300.
[2]Kim SS, Flannery B, Foppa IM, et al. Effects of prior season vaccination on current season vaccine effectiveness in the United States Flu Vaccine Effectiveness Network, 2012-2013 Through 2017-2018[J]. Clin Infect Dis, 2021, 73:497-505.
[3]Zhu L, Han Y, Lu J, et al. Evaluation of influenza vaccine effectiveness from 2021 to 2024: a Guangdong-based test-negative case-control study[J]. Vaccines (Basel), 2024, 13.doi:10.3390/vaccines13010004.
[4]Marchi S, Fallani E, Salvatore M, et al. The burden of influenza and the role of influenza vaccination in adults aged 50-64 years: a summary of available evidence[J]. Hum Vaccin Immunother, 2023, 19:2257048.doi:10.1080/21645515.2023.2257048.
[5]Sekiya T, Ohno M, Nomura N, et al. Selecting and using the appropriate influenza vaccine for each individual[J]. Viruses, 2021, 13.doi:10.3390/v13069071.
[6]Kim Y, Zheng X, Eschke K, et al. MCMV-based vaccine vectors expressing full-length viral proteins provide long-term humoral immune protection upon a single-shot vaccination[J]. Cell Mol Immunol, 2022, 19:234-244.
[7]Bliss CM, Freyn AW, Caniels TG, et al. A single-shot adenoviral vaccine provides hemagglutinin stalk-mediated protection against heterosubtypic influenza challenge in mice[J]. Mol Ther, 2022, 30:2024-2047.
[8]Trombetta CM, Marchi S, Manini I, et al. Challenges in the development of egg-independent vaccines for influenza[J]. Expert Rev Vaccines, 2019, 18:737-750.
[9]Wei CJ, Xu L, Kong WP, et al. Comparative efficacy of neutralizing antibodies elicited by recombinant hemagglutinin proteins from avian H5N1 influenza virus[J]. J Virol, 2008, 82:6200-6208.
[10]Cornelissen LA, de Vries RP, de Boer-Luijtze EA, et al. A single immunization with soluble recombinant trimeric hemagglutinin protects chickens against highly pathogenic avian influenza virus H5N1[J]. PLoS One, 2010, 5:e10645.doi:10.1371/journal.pone.0010645.
[11]Wiley DC, Skehel JJ. The structure and function of the hemagglutinin membrane glycoprotein of influenza virus[J]. Annu Rev Biochem, 1987, 56:365-394.
[12]Cox MM, Karl Anderson D. Production of a novel influenza vaccine using insect cells: protection against drifted strains[J]. Influenza Other Respir Viruses, 2007, 1:35-40.
[13]Bosch BJ, Bodewes R, de Vries RP, et al. Recombinant soluble, multimeric HA and NA exhibit distinctive types of protection against pandemic swine-origin 2009 A(H1N1) influenza virus infection in ferrets[J]. J Virol, 2010, 84:10366-10374.
[14]Weldon WC, Wang BZ, Martin MP, et al. Enhanced immunogenicity of stabilized trimeric soluble influenza hemagglutinin[J]. PLoS One, 2010, 5.doi:10.1371/journal.pone.0012466.
[15]金亚珉, 杨姣姣, 张婷, 等. B/Yamagata系流感病毒血凝素(HA)蛋白疫苗的筛选研究[J]. 现代生物医学进展, 2024, 24:2801-2804+2816.

可溶性流感病毒B/Victoria系血凝素重组蛋白的筛选

Screening of soluble hemagglutinin recombinant proteins for influenza virus B/Victoria

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