模拟肽聚糖表位的序列分析及其抗原性鉴定

发布时间：2018-07-17 01:37

【摘要】：目的:在线预测肽聚糖(PGN)模拟抗表原位并鉴定其抗原性。方法:利用抗PGN单克隆抗体从12线性噬菌体随机展示肽库中筛选模拟PGN表位的阳性克隆,对阳性克隆进行DNA测序和推导氨基酸序列,结合生物信息学分析阳性序列GRWxHxVxWAGL的抗原性以及T细胞表位,根据分析对阳性序列进行修饰与合成,ELISA法鉴定阳性序列的抗原性。结果:经对噬菌体12线性肽库的3轮筛选,夹心ELISA鉴定得到16个与抗PGN单克隆抗体结合的克隆,DNA测序并推导氨基酸序列,该序列与前期具有抗金黄色葡萄球菌活性序列No.31(ATWxHxLxSAGL)含有保守序列WxHx…AGL。在线分析(http://bio.dfci.harvard.edu/Tools/antigenic.pl,www.syfpeithi.de,http://www.darrenflower.info/mhcpred)表明,含保守序列的阳性序列GRWxHxVxWAGL包含与人和小鼠MHC结合表位(-WxHxVxW-),C端加入AGGS后具有T细胞表位(DNAstar),据此合成线性肽Biotin-GRWxHxVxWAGLAGGS(命名为SP39)。ELISA结果显示,SP39能与抗PGN单抗及抗S.aureus全菌多抗结合,PGN能抑制SP39与抗PGN单克隆抗体结合。结论:经噬菌体肽库筛选获得阳性序列GRWxHxVxWAGLAGGS,该序列可能模拟PGN抗原表位。
[Abstract]:Objective: to predict the antigenicity of peptidoglycan (PGN) in situ. Methods: the positive clones of PGN epitope were screened from 12 linear phage display peptide library using anti-PGN monoclonal antibody. The positive clones were sequenced and deduced amino acid sequences. Combined with bioinformatics to analyze the antigenicity and T cell epitope of GRWxHxVxWAGL, the positive sequence was modified and the antigenicity of the positive sequence was identified by synthetic Elisa. Results: after three rounds of screening of phage 12 linear peptide library, 16 clones were identified by sandwich Elisa, which combined with anti-PGN monoclonal antibody and deduced amino acid sequence. This sequence contains conserved sequence WxHx with previous antiStaphylococcus aureus active sequence No.31 (ATWxHxLxSAGL). AGL. The online analytics (http:% -bio.dfci.harvard.edur / tools / antigenic.plnn www.syfpeithi.deu httpw / r / www.darrenflower.info-mhcpred) shows that, The positive sequence GRWxHxVxWAGL containing conserved sequence contains T-cell epitope (DNAstar) after the C-terminal of human and mouse MHC binding epitope (-WxHxVxW-) is added to AGGS. According to this, the linear peptide Biotin-GRWxHxVxWAGLAGGS (named SP39). Elisa results show that SP39 can bind PGN with anti-PGN monoclonal antibody and anti-S.aureus polyclonal antibody. It can inhibit the binding of SP39 to anti PGN monoclonal antibody. Conclusion: the positive sequence GRWxHxVxWAGLAGGSs was obtained by phage peptide library screening, which may mimic the PGN epitope.
【作者单位】：江西省人民医院检验科;南方医科大学基础医学院免疫教研室;
【基金】：国家自然科学基金(No.31270980) 广东省医学科研基金(A2016533)资助
【分类号】：R392-33

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