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目的:预测及鉴定结核杆菌(Mycobacterium tuberculosis,Mtb)抗原Ag85C的HLA-A*0201限制性CD8+CTL表位,为基于表位的结核疫苗研究提供依据。方法:应用SYFPEITHI数据库预测结核杆菌抗原Ag85C序列中可能存在的HLA-A*0201限制性T细胞表位,利用T2细胞株分析各预测的抗原肽与HLA-A*0201分子的亲合力,选取高亲合力肽诱导特异性CTL细胞,检测各高亲合力肽刺激其特异性CTL细胞分泌的IFN-γ水平、在体外的CTL增殖反应以及细胞杀伤毒性,逐步鉴定出Ag85C的HLA-A*0201限制性CD8+CTL表位。结果:SYFPEITHI数据库从Ag85C序列中预测到14条能够结合HLA-A*0201分子的抗原肽,其中3个抗原肽(170～178 aa、317～325 aa和144～153 aa)显示与T2细胞上HLA-A*0201分子有高结合力,而抗原肽FLTREMPAWL(144～153 aa)能够诱导大多数HLA-A*0201阳性结核患者及PPD(+)健康志愿者产生特异性CTL细胞,并分泌大量的IFN-γ,能够诱导CTL体外发生增殖,能够产生特异性杀伤活性。结论:成功鉴定出抗原肽FLTREMPAWL(144～153 aa)结核杆菌抗原Ag85C的HLA-A*0201限制性CD8+CTL表位,可作为结核疫苗设计的候选表位,为进一步研发新型有效的抗结核疫苗奠定了基础。
OBJECTIVE: To predict and identify HLA-A * 0201-restricted CD8 + CTL epitopes of Mycobacterium tuberculosis (Mtb) antigen Ag85C and provide basis for epitope-based TB vaccine research. Methods: The HLA-A * 0201-restricted T cell epitopes in Mycobacterium tuberculosis antigen Ag85C sequence were predicted by SYFPEITHI database. The affinity of each predicted antigen peptide to HLA-A * 0201 molecule was analyzed by T2 cell line. The specific CTL cells were induced by avidin and the level of IFN-γ secreted by specific CTL cells stimulated by high avidity peptides was measured. CTL proliferation reaction and cytotoxicity in vitro were detected, and the HLA-A * 0201 limit of Ag85C was gradually identified Sex CD8 + CTL epitopes. RESULTS: The SYFPEITHI database predicted 14 antigen peptides capable of binding to HLA-A * 0201 from the Ag85C sequence, of which 3 antigenic peptides (170-178 aa, 317-325 aa and 144-153 aa) The HLA-A * 0201 molecule has a high binding capacity, whereas the antigen peptide FLTREMPAWL (144-153 aa) induces the production of specific CTL cells in most HLA-A * 0201-positive tuberculosis patients and healthy PPD (+) volunteers, Of IFN-γ, CTL can induce proliferation in vitro, can produce specific killing activity. CONCLUSION: The HLA-A * 0201 restricted CD8 + CTL epitope of Mycobacterium tuberculosis antigen Ag85C was successfully identified as candidate epitope of tuberculosis vaccine. To further develop a new and effective anti-TB Vaccine laid the foundation.