胃癌相关抗原MG7-Ag模拟肽表位的筛选及测序分析

目的　从噬菌体呈现的随机肽库中筛选胃癌单克隆抗体MG7所识别的胃癌相关抗原的短肽模拟表位 ,为进一步研究胃癌相关抗原与单抗结合的结构基序奠定基础。方法　用胃癌单克隆抗体MG7对呈现于噬菌体衣壳蛋白pⅧ上的 2个九肽库分别进行亲和富集和免疫筛选 ;阳性克隆进一步用荧光标记和硝酸纤维素膜斑点印迹法证实其结合活性 ;经随机抽取部分阳性克隆进行DNA序列分析 ,推导出相应噬菌体呈现肽的氨基酸序列 ,通过序列比较分析相对保守的表位信息 ;运用HLA分子结合分析软件预测已测序短肽与HLA分子结合的可能性。结果　经反复多轮筛选和结合活性检测 ,从pⅧ和pⅧ cys 2个噬菌体肽库中分别得到了 12和 30个阳性克隆 ;通过对部分阳性克隆进行测序分析 ,推导相应的随机肽序列和序列比较分析 ,得到具有相对保守性的表位信息如PLX0 2 S、SAVR、XRMX、YARN等。经计算机预测分析 ,发现它们可与HLA多个分子结合。结论　PLX0 2 S、SAVR、XRMX、YARN等有可能为胃癌单克隆抗体MG7所识别的表位基序。

Mimicepitope recognized by monoclonal antibody MG7 against gastric cancer through screeningphage displayed random peptide library

OBJECTIVE: To get mimic peptide epitopes that could be recognized by a monoclonal antibody against gastric cancer named MG7 from random peptide library displayed by phage, and to provide useful information for further study of the interaction between antigen and antibody. METHODS: Through affinity enrichment and immunoscreening of two phage-displayed non-apeptide libraries constructed in pVIII with MG7 MAb separately, several positive phages were obtained. Fluorescence labeling and dot blot were carried out for further identification of their binding activities. Then, some of the positive phages were sequenced and their corresponding peptide sequence was deduced according to their DNA sequence. Finally, HLA binding prediction software was applied for HLA binding analysis. RESULTS: Based on several rounds of screening and binding activity detection, we got twelve and thirty positive phage clones respectively from two libraries. Through DNA sequencing, peptide deducing and sequence aligning analysis of the positive phages, some preserved epitope information such as PLX(0 - 2)S, SAVR, XRMX and YARN were obtained. The prediction using HLA binding analysis software showed that most of the sequenced peptide had the potentiality to bind with HLA molecules. CONCLUSION: PLX(0 - 2)S, SAVR, XRMX and YARN may be some of the motifs which could be recognized by monoclonal antibody MG7.