应用噬菌体肽库研究银屑病相关的角蛋白K14,K17模拟表位(英文)

Screening for the mimetic homologous oligopeptides of keratins 14 and 17 related with psoriasis from phage random peptide library

目的 获得与银屑病相关的角蛋白 K1 4和 K1 7同源序列的模拟表位 ,评估含有此基序的短肽与银屑病发病的关系 .方法 将 1株抗角蛋白 K1 4和 K1 7同源序列的单克隆抗体 (m Ab) 5 G5经亲和层析纯化后进行生物素标记 ,对噬菌体递呈的随机 6肽库进行 3轮淘洗并进行 EL ISA检测 .挑取1 0个阳性克隆进行 DNA测序 ,分析所获数据 ,并进行竞争阻断实验 .结果 氨基酸序列分析表明模拟短肽的基序为 VL(x) AG,角蛋白 K1 4、K1 7的同源序列及链球菌 M蛋白均含有此基序 .携有这些短肽的噬菌体可与 m Ab5 G5特异结合 ,并可阻断单抗与角蛋白反应 .结论 含有此基序的短肽可以模拟 m Ab5 G5识别的与银屑病相关的角蛋白 K1 4和 K1 7同源序列的抗原表位 ,为银屑病特异性短肽的研究提供了一个新途径 。

AIM To acquire the mimetic homologous oligopeptides of human epidermal keratins 14 and 17 related with psoriasis and evaluate the effect of the oligopeptides on the pathogenesis of psoriasis. METHODS The mAb 5G5 recognizing the common epitope of human epidermal keratin K14 and K17 was purified by HiTrap Protein G affinity column, and was biotinylated by the biotin ester. A 6 mer phage random peptide library was biopanned for 3 cycles, then positive clones were identified by ELISA and DNA were extracted for sequencing. RESULTS Amino acid sequence analysis showed that 10 positive clones selected randomly had the consensus Amino acid sequence (motif) VL(x)AG. The motif VL(x)AG could be detected in the homologous amino acid sequence of keratins 14 and 17, and streptococcal M protein contained the motif too. The phages of positive clones reacted with mAb 5G5 specifically and prevented the interaction between mAb 5G5 and keratins with dose dependent effects. CONCLUSION The motif could mimic the common epitope on human epidermal keratins 14,17 and streptococcal M protein, perhaps the research could provide a new approach for the discovery of psoriatic specific peptide epitope and the preparation of peptide vaccine for psoriasis therapy.