H5Nx禽流感病毒交叉反应性CD4+ T细胞表位预测

doi:10.13560/j.cnki.biotech.bull.1985.2023-1006

摘要/Abstract

摘要：

【目的】评估预先存在的H5Nx交叉反应性免疫记忆，为人类H5禽流感病毒的防治和广谱疫苗的研发提供理论数据。【方法】利用生物信息学方法筛选人源H5Nx禽流感病毒与季节性流感病毒的共同保守的交叉反应性CD4⁺ T细胞表位，进一步分析保守表位嵌套CD8⁺ T细胞表位和B细胞表位情况以及在中国人群中的覆盖率。【结果】92.3%（513/555）的H5Nx CD4⁺ T细胞表位在季节性甲型流感病毒中具有交叉反应性，其中172个表位嵌套CD8⁺ T细胞表位和5个表位嵌套B细胞表位。这些表位与相应HLA-DRB1等位基因在全世界人群中的覆盖率为88.65%。【结论】由于反复接触季节性甲型流感病毒，人群中存在一定水平的的H5Nx交叉反应性免疫记忆，能够为H5Nx感染提供部分保护。

关键词: H5Nx流感病毒, 共同保守, 交叉反应性, CD4⁺ T细胞表位, 嵌套表位

Abstract:

【Objective】To evaluate preexisting H5Nx cross-reactivity immune memory and to provide theoretical data for the prevention and treatment of human H5 avian influenza viruses and the development of broad-spectrum vaccines.【Method】Bioinformatics methods were used to screen the common conserved cross-reactivity CD4⁺ T cell epitopes of human H5Nx avian influenza viruses and seasonal influenza viruses, and further analysis of the nested CD8⁺ T cell epitopes and B cell epitopes of conserved epitopes and their coverage among the Chinese population.【Result】92.3%（513/555）of H5Nx CD4⁺ T cell epitopes were cross-reactivity in seasonal influenza A viruses, of which 172 epitopes nested CD8⁺ T cell epitopes and 5 epitopes nested B cell epitopes. These epitopes and the corresponding HLA-DRB1 alleles have a strong coverage rate of 88.65% in the world population.【Conclusion】Due to repeated exposure to seasonal influenza A viruses, there is a certain level of H5Nx cross-reactivity immune memory in the population that may provide partial protection from H5Nx infection.

Key words: H5Nx influenza virus, co-conserved, cross-reactivity, CD4⁺ T-cell epitopes, nested epitopes

谢爽, 陈瑶, 黄俊琼. H5Nx禽流感病毒交叉反应性CD4⁺ T细胞表位预测[J]. 生物技术通报, 2024, 40(5): 300-309.

XIE Shuang, CHEN Yao, HUANG Jun-qiong. Prediction of H5Nx Avian Influenza Virus Cross-reactivity CD4⁺ T-cell Epitopes[J]. Biotechnology Bulletin, 2024, 40(5): 300-309.

图/表 8

表1 sH1N1、pH1N1、H3N2、H5N1、H5N6和H5N8病毒蛋白质序列分析数据集

Table 1 Protein sequence analysis datasets of sH1N1, pH-1N1, H3N2, H5N1, H5N6 and H5N8 virus subtypes

蛋白片段Protein segment	蛋白序列数 Protein sequence number
蛋白片段Protein segment	sH1N1	pH1N1	H3N2	H5N1	H5N6	H5N8	Total
HA	1994	43360	62270	517	37	1	108179
NA	2803	39867	56908	477	37	1	100093
M1	1645	28497	44559	394	37	1	75133
M2	1645	28496	44569	404	37	1	75152
NS1	872	22279	33622	391	37	1	57202
NEP	872	22279	33624	389	37	1	57202
NP	881	22047	35759	384	37	1	59109
PA	877	22522	35229	386	37	1	59052
PB1	871	20601	34524	371	37	1	56405
PB2	900	21837	34855	371	37	1	58001

表2 A/Astrakhan/3212/2020（H5N8）与sH1N1、pH1N1和H3N2病毒氨基酸序列同源性

Table 2 Amino acid sequence identity of A/Astrakhan/3212/2020（H5N8）with sH1N1, pH1N1 and H3N2 virus subtypes

蛋白片段 Protein segment	氨基酸序列同源性 Amino acid sequence identity/%
蛋白片段 Protein segment	sH1N1	pH1N1	H3N2	H5N1	H5N6
HA	50.5-56.2	47.1-55.4	32.8-55.6	88.4-94.2	92.5-99.3
NA	16.7-39.4	32.6-39.7	19.1-37.0	38.2-43.1	18.7-22.6
M1	92.2-96.8	91.3-95.5	90.0-96.8	92.5-97.1	93.4-100.0
M2	73.7-91.4	80.7-91.4	72.4-93.6	79.1-92.2	80.7-100.0
NS1	76.2-86.2	72.3-80.7	72.6-86.7	80.6-89.8	58.5-93.2
NEP	85.8-93.2	79.9-89.6	82.9-91.4	82.5-92.3	76.9-96.6
NP	90.5-96.3	90.3-94.5	88.5-95.7	96.4-98.4	93.4-97.6
PA	91.6-96.7	93.5-97.0	92.2-97.0	94.5-98.3	95.1-98.0
PB1	93.9-97.3	93.9-96.4	93.7-98.0	96.0-98.3	96.0-98.5
PB2	93.5-96.1	92.9-96.9	87.7-97.2	95.0-97.9	95.7-98.4

表3 A/Astrakhan/3212/2020（H5N8）的CD4+ T细胞预测表位

Table 3 Predicted CD4+ T-cell epitopes of A/Astrakhan/3212/2020（H5N8）

蛋白片段 Protein segment	（A）预测的CD4⁺ T细胞表位（A）Predicted CD4⁺T-cell epitopes	（B）保守表位（%=B/A100）（B）Conserved predicted epitopes（% = B/A100）	（C）交叉反应性表位（%=C/B100）（C）Cross-reactivity epitopes（%= C/B100）	已验证的表位（来源：IEDB，免疫表位数据库） Experimentally defined epitopes/% (Source: IEDB, immune epitope database)
蛋白片段 Protein segment	（A）预测的CD4⁺ T细胞表位（A）Predicted CD4⁺T-cell epitopes			（D）CD4⁺ T细胞表位（%=D/C100）（D）CD4⁺ T-cell epitopes（%=D/C100）	（E）CD8⁺ T细胞表位（%=E/D100）（E）CD8⁺ T-cell epitopes（%=E/D100）	（F）B细胞表位（%=F/E100）（F）B-cell epitopes（%=F/E100）
HA	102	24	7	7	7	2
NA	56	0	0	0	0	0
M1	77	47	47	40	29	3
M2	8	0	0	0	0	0
NS1	46	4	4	4	0	0
NEP	29	8	6	4	4	0
NP	118	98	75	57	49	0
PA	122	104	104	32	21	0
PB1	158	138	138	63	42	0
PB2	179	132	132	27	20	0
Total	895	555（62.0）	513（92.4）	234（45.6）	172（73.5）	5（2.9）

表4 H5Nx的CD4+ T细胞表位与季节性甲型流感病毒的交叉反应性

Table 4 Cross-reactivity between CD4+ T-cell epitopes of H5Nx and seasonal IAV

蛋白片段 Protein segment	H5Nx预测表位 H5Nx predicted epitopes	与H5Nx的交叉表位 Cross-epitopes with H5Nx
蛋白片段 Protein segment	H5Nx预测表位 H5Nx predicted epitopes	sH1N1	pH1N1	H3N2
HA head	8	0	0	0
HA stem	16	7	4	0
NA	0	0	0	0
M1	47	41	45	40
M2	0	0	0	0
NS1	4	4	4	4
NEP	8	6	6	6
NP	98	61	66	52
PA	104	95	96	93
PB1	138	129	122	138
PB2	132	120	128	121
Total	555	463	471	454

图1 A/Astrakhan/3212/2020（H5N8）HA和M1的拉试图

Fig. 1 Ramachandran plot of A/Astrakhan/3212/2020（H5N8）HA and M1

图2 A/Astrakhan/3212/2020（H5N8）HA-A和M1-D嵌套表位的3D模拟 A: HA头部和茎部分别用浅灰色和深灰色标记；绿色标记CD4+ T细胞表位，浅蓝色标记嵌套CD8+ T细胞表位，紫色标记嵌套表位B 细胞表位； D: M蛋白其余氨基酸用棕色标记；B、C和 E、F: 分别是与CD8+ T细胞和 B 细胞表位嵌套的 CD4+ T 细胞表位的卡通和棒表示，虚线代表氢键

Fig. 2 Locating nested epitopes in 3D model of A/Astrakhan/3212/2020（H5N8）HA-A and M1-D A marks the HA head and stem domains in light gray and dark gray respectively; green marks CD4+ T-cell epitopes, light blue marks nested CD8+ T-cell epitopes, and purple marks nested B-cell epitopes; D marks other amino acid residues of M1 in brown; B, C and E, F are respectively cartoon and sticks representation of CD4+ T-cell epitopes nested with both CD8+ T-cell and B-cell epitopes. The dashed lines represent hydrogen bonds

图3 IAV交叉反应性CD4+ T细胞表位的HLA-DRB1等位基因限制性

Fig. 3 HLA-DRB1 allele-restriction of IAV cross-reactivity CD4+ T-cell epitopes

图4 IAV交叉反应性CD4+ T细胞表位的人口覆盖率 A：所有种族群体；B：H5N8 中的8种蛋白质。已确定的交叉反应性 CD4+ T 细胞表位提供了广泛的人群覆盖范围。根据每个 HLA II 类限制 DRB1 等位基因的结合数据，计算理论群体覆盖率。显示了可能的表位-HLA 等位基因组合的数量作为每个种族群体的分数（%）和每个蛋白质（%）的函数

Fig. 4 Population coverage of identified cross-reactivity CD4+ T-cell epitopes A: All ethnicities groups. B: Eight proteins in H5N8. The identified cross-reactivity CD4+ T-cell epitopes provide broad population coverage. Based on the binding data for each HLA class II-restricted DRB1 alleles, theoretical population coverage was calculated. The number of possible epitope-HLA allele combinations as a function of the fraction of each ethnic population（%）and each protein（%）is shown

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