Epitope Mapping Using Ribosome Display in a Reconstituted Cell-Free Protein Synthesis System
Epitope Mapping Using Ribosome Display in a Reconstituted Cell-Free Protein Synthesis System
复制标题
DOI：
10.1093/jb/mvp027
复制
发表时间：
2009-05-01
影响因子：
2.7
通讯作者：
Ueda, Takuya
中科院分区：
文献类型：
--
作者：
Osada, Eriko;Shimizu, Yoshihiro;Ueda, Takuya
核糖体展示技术是筛选配体结合肽或蛋白质的有力手段。我们在这里证明，核糖体展示使用重建的无细胞蛋白质合成系统可以应用于单克隆抗体（mAb）的表位作图。利用该技术，我们从随机肽库中筛选出与三种单克隆抗体特异性结合的肽。当针对抗FLAG M2抗体进行选择时，所选肽含有先前表征的共有表位，表明该方法可用于表位作图。当对两种抗-Catenin（抗-Cat）单克隆抗体进行选择时，所选肽具有-Cat的部分肽序列的同源性。Western blot分析表明，这些推定的表位具有相应的单克隆抗体和-Cat突变体的亲和力，缺乏这些区域不结合抗体，表明我们正确地映射了这些单克隆抗体的表位。本研究为单克隆抗体表位的快速鉴定提供了一种方法。
Ribosome display is a powerful technology for selecting ligand-binding peptides or proteins. We demonstrate here that the ribosome display using the reconstituted cell-free protein synthesis system can be applied for the epitope mapping of monoclonal antibodies (mAbs). Using this technology, we selected peptides that specifically bind to three mAbs from random peptide library. When selection was performed against the anti-FLAG M2 antibody, selected peptides contained previously characterized consensus epitope, indicating that the methodology can be applied for the epitope mapping. When the selection was carried out against two anti--Catenin (anti--Cat) mAbs, selected peptides had a homology for the partial peptide sequences of -Cat. Western blot analysis showed that these putative epitopes had affinity for the corresponding mAbs and -Cat mutants that lack these regions did not bind to the antibodies, indicating we correctly mapped the epitope for these mAbs. The study shown here provides a way for the quick identification of the epitope of mAbs.