应用噬菌体展示肽库技术淘选大鼠CCR5膜外第一、二胞外环特异性结合的活性拮抗肽与初步鉴定

Panning and identification of antagonistic active peptides specifically binding to the first and second extracellular membrane loops of rat CCR5 by technique of phage display peptide library

目的：利用噬菌体展示肽库技术淘选与大鼠CC趋化因子受体5（CCR5）膜外第一、二胞外环特异性结合的短肽,并鉴定其与CCR5的结合能力。方法：在蛋白质数据库中查得大鼠CCR5第一、二胞外环的氨基酸序列,合成相应的线性短肽作为淘选的靶分子,利用噬菌体展示7肽文库进行3~4轮淘选,用ELISA法鉴定所选肽与靶分子的结合,并测定其与浓度的关系。结果：与CCR5第一、二胞外环特异性结合的噬菌体展示的短肽序列分别为GHWKVWL和HYIDFRW,ELISA鉴定呈阳性反应,且短肽与靶分子的结合具有浓度依赖性和可饱和性。结论：利用噬菌体展示技术成功获得了2条CCR5特异性结合的短肽,并在体外证明其可与CCR5第一、二胞外环具有结合能力。

AIM： To pan the active peptides which specifically bound to the first and second extracellular membrane loops of rat CC chemokine receptor 5（ CCR5）. METHODS： The technique of phage display peptide library was used and binding ability of the peptides was identified. The amino acid sequences of the first and second extracellular loops of rat CCR5 were searched in the protein database and chemically synthesized corresponding linear peptides were used as targets in the biopanning. After 3 to 4 rounds of screening with Ph. D.TM-7 Phage Display Peptide Library were performed,the specific phages were collected and primarily identified by ELISA. RESULTS： The sequences of the peptides displayed on the selected phages were GHWKVWL and HYIDFRW,both of them exhibited positive in phage binding ELISA and the binding to phages and targets were concentration dependent and saturable. CONCLUSION： Two antagonistic active peptides specifically binding to CCR5 were successfully obtained by the technique of phage display peptide library,and the binding ability to the first and second extracellular membrane loops of rat CCR5 were proved in vitro.