噬菌体展示的小肽序列差异对肿瘤坏死因子-α诱导的细胞毒作用的影响  

Affect of phage-displayed small peptide sequences on TNF-α-mediated cytotoxicity in vitro

作　　者：刘新平[1] 张晓光[1] 韩炯[1] 李福洋[1] 王吉村[1] 药立波[1] 

机构地区：[1]第四军医大学基础部生物化学与分子生物学教研室,西安710032

出　　处：《中华风湿病学杂志》2001年第3期142-144,共3页Chinese Journal of Rheumatology

基　　金：国家自然科学基金;杰出青年基金!资助(3 982 5 113 )

摘　　要：目的　从噬菌体随机多肽库中筛选出与肿瘤坏死因子 α(TNF α)特异结合 ,并能阻断其生物活性的小肽 ,为建立一种新型的抗TNF α多肽药物奠定基础。方法　用突变体TNF α和天然型TNF α分别作为靶蛋白 ,用噬菌体随机多肽库进行多次淘筛 ,经序列测定及体外酶联免疫吸附法(ELISA)实验选出候选多肽 ,进行抗TNF α诱导的L92 9细胞毒作用。结果　筛选到的二个序列不同但有一定相关性的候选小肽 ,其中由天然型TNF α作为靶蛋白筛选出的 12肽具有抗TNF α诱导的L92 9细胞毒作用。结论　这些序列可以做为合成小肽TNFObjective By using phage displayed 12 peptide libraries to select antagonists to TNF α which could inhibit TNF α induced cytotoxicity in vitro .Methods A random 12 mer peptide library was screened with 1 ml rhTNF α (100 μg),according to biopanning procedure of New England Biolabs Inc.Combinant human TNF α (2 2×10 8 U/ml) were purchased from Biotechnology Centre.The mouse fibroblast cell L929 was used to measure TNF α mediated cytotoxicity.DNA was sequenced on an ABI PRISM automated 377 DNA sequencer.Results Randomly selected phage clones after the third biopanning were tested for their special binding with TNF α in ELISA.The 21 positive clones were sequenced with an ABI PRISM Dye terminator cycle sequencing reaction kit.From the 21 clones tested,one (11),consistently inhibited the TNF α induced cytotoxicity on L929 cells.The inhibition was dose dependent at concentration of TNF α giving 38 2%.No inhibition was observed with a control phage.Conclusion These sequences may take for the basis of sythesis of small peptide antagonist to TNF α.

关 键 词：肿瘤坏死因子 肽库 类风湿性关节炎 

分 类 号：R593.22[医药卫生—内科学]