HIV-1gp41 C-螺旋模拟位的筛选和鉴定  

Screening and identification of HIV-1 gp41 C-helix mimotopes displayed on phages

作　　者：刘北一[1] 朱平[1] 韩强涛[1] 姜世勃 富宁[1] 

机构地区：[1]第一军医大学基础部免疫学教研室,广州510515 [2]纽约血液中心LFK研究所,ny10021

出　　处：《中国免疫学杂志》2004年第9期633-636,共4页Chinese Journal of Immunology

基　　金：国家自然科学基金海外合作基金资助项目 ( 3 0 0 2 80 2 1)

摘　　要：目的 :筛选可作为小分子先导化合物的HIV 1gp4 1C螺旋模拟位 ,为开发抗HIV 1早期感染的小分子药物奠定基础。方法 :以源于HIV 1gp4 1N端的肽N36为靶 ,对噬菌体环七肽库进行亲和筛选。利用ELISA鉴定噬菌体阳性克隆并对其进行DNA序列分析。结果 :3轮筛选后随机挑取 2 6个噬菌体克隆 ,ELISA鉴定表明有 16个克隆可与N36结合 ,将其中 10个克隆DNA测序并推导氨基酸序列 ,每一克隆至少含有 2个疏水氨基酸 ,其中 9个克隆均有WW ,7个克隆有WWH保守序列。挑选阳性噬菌体克隆No 8进一步鉴定 ,游离N36肽阻断No 8克隆与固相化N36结合 ,C34肽竞争抑制N36肽与No 8克隆结合 (IC50 为 12 5 μg ml)。 结论 :表达WW保守序列的肽模拟HIV 1gp4 1C螺旋与N螺旋结合 ,该结果为设计针对HIV介导融膜的抑制剂提供技术支持。Objective:To find small molecular leads for inhibition on early stage of HIV infection by identification and characterization of the HIV-1 gp41 C-helix mimotopes.Methods:For identification of the gp41 C-helix mimotopes,C7C phage display peptide library was biopanning by using a synthetic peptide N36 which was derived from the gp41 N-helix as target.After three rounds of screening,positive phage clones were identified by ELISA and sequenced.Results:16 of 26 phage clones were identified to bind with peptide N36,and 10 of them were sequenced.Every clone of ten clones contains at least two hydrophobic residues,which may dock into the hydrophobic pocket in the gp41 N-helix domain.9 of the 10 clones have a conservative sequence WW,which may mimic the W628 and W631 in C-helix to interact with the hydrophobic residues in the gp41 pocket.One clone expressing the conservative sequence named clone No.8(CYWWHRLHC) was selected for characterization.The binding between the clone No.8 and N36 was blocked by free peptide N36.And the binding between clone No.8 and peptide N36 was inhibited by peptide C34(IC 50=12.5 μg/ml).Conclusion:The short circular peptides displayed on phages containing WW residues may mimic the conformational epitope of the HIV-1 gp41 C-helix to interact with the N-helix.This information may be useful for design of HIV-1 fusion inhibitors.

关 键 词：HIV-1 GP41 N-螺旋 C-螺旋 模拟位 噬菌体肽库 

分 类 号：R392-33[医药卫生—免疫学]