通过使用固定在聚乙二醇胺树脂上的特异性配体进行亲和纯化来鉴定小分子结合分子。

Identification of small molecule binding molecules by affinity purification using a specific ligand immobilized on PEGA resin.

作者信息

Kuramochi Kouji, Miyano Yuka, Enomoto Yoshihiro, Takeuchi Ryo, Ishi Kazutomo, Takakusagi Yoichi, Saitoh Takeki, Fukudome Keishi, Manita Daisuke, Takeda Yoshifumi, Kobayashi Susumu, Sakaguchi Kengo, Sugawara Fumio

机构信息

Department of Applied Biological Science, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.

出版信息

Bioconjug Chem. 2008 Dec;19(12):2417-26. doi: 10.1021/bc8002716.

DOI:10.1021/bc8002716

PMID:19035789

Abstract

We investigated the application of resins used in solid-phase synthesis for affinity purification. A synthetic ligand for FK506-binding protein 12 (SLF) was immobilized on various resins, and the binding assays between the SLF-immobilized resins and FK506-binding protein 12 (FKBP12) were performed. Of the resins tested in this study, PEGA resin was the most effective for isolating FKBP12. This matrix enabled the isolation of FKBP12 from a cell lysate, and the identification of SLF-binding peptides from a phage cDNA library. We confirmed the interaction between SLF and these peptides using a cuvette type quartz crystal microbalance (QCM) apparatus. Our study suggests that PEGA resin has great potential as a tool not only for the purification and identification of small-molecule binding proteins but also for the selection of peptides that recognize target molecules.

摘要

我们研究了用于亲和纯化的固相合成树脂的应用。将一种用于FK506结合蛋白12（SLF）的合成配体固定在各种树脂上，并进行了固定有SLF的树脂与FK506结合蛋白12（FKBP12）之间的结合测定。在本研究中测试的树脂中，PEGA树脂在分离FKBP12方面最有效。这种基质能够从细胞裂解物中分离出FKBP12，并从噬菌体cDNA文库中鉴定出与SLF结合的肽段。我们使用比色皿型石英晶体微天平（QCM）仪器证实了SLF与这些肽段之间的相互作用。我们的研究表明，PEGA树脂不仅作为一种工具在纯化和鉴定小分子结合蛋白方面具有巨大潜力，而且在选择识别靶分子的肽段方面也具有巨大潜力。