烟曲霉抗原Asp f16 HLA-A-* 0201限制性的CD8-+ CTL抗原表位生物信息学预测与实验室鉴定

中国真菌学杂志　 2011, Vol. 6 　Issue (2): 87-92.

烟曲霉抗原Asp f16 HLA-A-* 0201限制性的CD8-+ CTL抗原表位生物信息学预测与实验室鉴定

赵作涛¹, 孙铮¹, 万喆¹, 朱平², 李若瑜¹

1. 北京大学第一医院皮肤性病科, 北京, 100034;
2. 北京大学真菌和真菌病研究中心, 北京, 100034;
3. 北京大学第一医院血液科, 北京, 100034

收稿日期:2010-12-31 出版日期:2011-04-28 发布日期:2011-04-28
通讯作者: 李若瑜,E-mail:lrymm@medmail.com.cn E-mail:lrymm@medmail.com.cn
作者简介:赵作涛,男(汉族),博士,助理研究员.E-mail:zhaozuotao@yahoo.com
基金资助:
国家自然科学基金青年基金(30800041)

Prediction and identification of new HLA-A-* 0201-restricted CD8-+ CTL epitopes from Aspergillusfumigatus antigen Asp f16

ZHAO Zuo-tao¹, SUN Zheng¹, WAN Zhe¹, ZHU Ping², LI Ruo-yu¹

1. Research Center for Medical Mycology, Beijing, 100034, China;
2. Department of Dermatology, Beijing, 100034, China;
3. Department of Hematology, First Hospital, Peking University, Beijing, 100034, China

Received:2010-12-31 Online:2011-04-28 Published:2011-04-28

摘要/Abstract

摘要： 目的预测与鉴定烟曲霉抗原Asp f16的HLA-A*0201限制性CD8+细胞毒性T细胞(CTL)抗原表位。方法以国人常见的HLA-A*0201位点为靶点,依据生物信息学软件扫描烟曲霉特异性抗原Asp f16的全部427个氨基酸序列。使用HLA-A*0201转基因小鼠制备骨髓来源的树突状细胞(DC)和CTL。流式细胞仪技术检测DC表面MHC II类抗原,CD80,CD86和CD11c的表达来验证其是否成熟。ELISPOT试验检测烟曲霉抗原多肽特异性CTL产生的细胞因子IFN-γ。四聚体(Tetramer)试验证实烟曲霉特异性CTL与抗原肽,HLA-A*0201分子复合体的亲和性。结果根据与MHC I类分子结合的半衰期评分,选择了3个HLA-A*0201限制性抗原表位。流式细胞仪分析示成熟DC高表达HLA II类抗原,CD80,CD86和CD11c。Tetramer试验证实烟曲霉特异性T细胞受体与抗原肽,HLA-A*0201分子复合体的高亲和性。ELISPOT实验结果表明烟曲霉抗原肽体外可以活化CD8+CTL,被负载了抗原肽的DC刺激活化后可以产生IFN-γ。结论本研究成功鉴定烟曲霉抗原Asp f16的HLA-A*0201限制性CD8+CTL表位,可作为疫苗设计的候选表位,为进一步研发新型抗烟曲霉疫苗提供参考。

关键词: 烟曲霉, Asp f16, 抗原表位, 树突状细胞, 细胞毒性T细胞

Abstract: Objective To predict and identify potential HLA-A*0201-restricted CD8+ cytotoxic T lymphocyte(CTL) epitopes in Aspergillus fumigatus antigen Asp f16.Methods A total of 427aa Asp f16 amino acid sequences were scanned using HLA peptide binding prediction software on the target of HLA-A*0201.Bone-marrow-derived dendritic cells(DC) and peptide specific CTLs were prepared in HLA A*0201 transgenic mice.Expressions of MHC class II antigen,CD80,CD86,and CD11c were analyzed by flow cytometry.IFN-γ produced by Asp f16 peptides specific CTLs was detected by ELISPOT assay.Tetramer assays were performed to investigate the affinity of the CD8+ CTLs to the Asp f16 peptides complexed HLA-A*0201.Results Three potential HLA-A*0201 binding 9-mer peptides were selected based on their estimated dissociationhalf-life from MHC class I molecular.DCshighly expressed MHC class II antigen,CD80,CD86 and CD11c.Strong affinity of the TCR to the peptide/MHC complex was demonstrated by tetramer staining.Asp f16 peptides specific CD8+ CTL actively produced IFN-γ after exposure to peptide-loaded DCs.Conclusions Three HLA-A*0201-restricted CD8+ CTL epitopes of Asp f16 successfully identified might be candidates for anti Apergillus fumigatus vaccine designing.

Key words: Aspergillus fumigatus, Asp f16, antigen epitope, dendritic cell, cytotoxic T lymphocyte

中图分类号:

R379.6

赵作涛, 孙铮, 万喆, 朱平, 李若瑜. 烟曲霉抗原Asp f16 HLA-A-* 0201限制性的CD8-+ CTL抗原表位生物信息学预测与实验室鉴定[J]. 中国真菌学杂志, 2011, 6(2): 87-92.

ZHAO Zuo-tao, SUN Zheng, WAN Zhe, ZHU Ping, LI Ruo-yu. Prediction and identification of new HLA-A-* 0201-restricted CD8-+ CTL epitopes from Aspergillusfumigatus antigen Asp f16[J]. Chinese Journal of Mycology, 2011, 6(2): 87-92.

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