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目的利用噬菌体12肽库筛选能与EB病毒(EBV)LMP1 CTAR23蛋白高效结合的短肽。方法利用亲和层析柱加酶切的方法纯化CTAR23蛋白,并用其筛选噬菌体12肽库。通过夹心ELISA法分析噬菌体克隆与CTAR23蛋白的亲和力。通过硝酸纤维膜斑点印迹法进行阳性克隆鉴定,并对阳性克隆进行测序及序列分析。结果对12肽库进行3轮筛选后,噬菌体克隆具有良好的富集效果。ELISA提示筛选出来的噬菌体克隆能与CTAR23蛋白高效结合。硝酸纤维膜斑点印迹法证实阳性率达到100%,测序结果显示噬菌体上的短肽有共同序列HVSLHKHYPTAS。结论噬菌体短肽HVSLHKHYPTAS为研究LMP1致瘤机理、开发拮抗LMP1蛋白的小分子短肽类药物奠定坚实的基础。
OBJECTIVE: To screen short peptide peptides that bind efficiently to Epstein-Barr virus (EBV) LMP1 CTAR23 protein using the phage 12 peptide library. Methods The CTAR23 protein was purified by affinity chromatography and enzyme digestion, and was used to screen phage 12 peptide library. The affinity of phage clones to the CTAR23 protein was analyzed by sandwich ELISA. The positive clones were identified by dot blotting with nitrocellulose membrane, and the positive clones were sequenced and sequenced. Results After three rounds of screening of 12 peptide libraries, the phage clones had a good enrichment effect. ELISA suggested that the selected phage clones could bind efficiently to CTAR23 protein. The positive rate was confirmed by dot blotting with nitrocellulose membrane. The sequencing results showed that the short peptide on phage had the common sequence HVSLHKHYPTAS. Conclusion The phage short peptide HVSLHKHYPTAS lays a firm foundation for the study of LMP1 tumorigenicity mechanism and the development of small molecule short peptides that antagonize the LMP1 protein.