隐球菌抗原MP98的HLA-A*0201限制性CD8+CTL表位的预测及鉴定

Prediction and Identification of HLA-A*0201 Restricted CD8+CTL Epitopes in Cryptococcus Neoformans MP98

目的鉴定隐球菌抗原MP98中的HLA-A*0201限制性CD8+CTL表位。方法应用数据库SYFPEITHI预测隐球菌MP98中可能存在的HLA-A*0201限制性CD8+CTL表位,经流式细胞术分析各抗原肽与HLA-A*0201的亲合力,经时间分辨荧光法检测外周血单个核细胞(PBMCs)对各抗原肽产生的增殖反应,经细胞毒性实验研究各抗原肽诱导的特异性T细胞的细胞毒杀伤活性,逐步鉴定MP98的HLA-A*0201限制性CD8+CTL表位。结果位于MP98氨基酸序列肽5(436-444aa)与HLA-A*0201分子具有较高的亲合力,并都能刺激HLA-A*0201阳性个体者PBMC增殖,并诱导产生具有特异性杀伤活性的CTL。结论肽5GMFDGLSGV(436-444aa)是MP98上HLA-A*0201限制性CD8+CTL的优势表位,可作为隐球菌疫苗设计的候选表位。

Objective To identify HLA-A ＊ 0201 restricted CD8＋CTL epitopes in Cryptococcus neoformans MP98. Methods Firstly,database SYFPEITHI was applied to predict HLA-A ＊ 0201 restricted CD8＋CTL epitopes in Cryptococcus neoformans MP98. Secondly,affinity and stability of the predicted peptides with T2 cell were assayed by flow cytometry. Thirdly,cells proliferation and cytotoxicity induced by peptides were investigated by DELFIA cell proliferation assay and DELFIA EUTDA cytotoxicity test respectively. Results Peptide 5 （GMFDGLSGV,436-444aa）had high affinity with HLA-A ＊ 0201 molecules,stimulated PBMCs from healthy donors to proliferate and induce antigen specific CTLs. Only pep6 could give a positive proliferation response and specific cytotoxicity. Conclusion Pep 5 GMFDGLSGV（436-444aa）is HLA-A ＊ 0201 restricted CDS＋CTL epitope of Cryptococcus neoformans MP98,which could be the candidate for peptide- based vaccine.