Creation and biochemical analysis of a broad-specific claudin binder.
Creation and biochemical analysis of a broad-specific claudin binder.
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DOI：
10.1016/j.biomaterials.2012.01.017
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发表时间：
2012-04
期刊：
影响因子：
14
通讯作者：
A. Takahashi;Yumiko Saito;M. Kondoh;K. Matsushita;S. Krug;Hidehiko Suzuki;H. Tsujino;Xiangru Li;H. Aoyama;Koji Matsuhisa;T. Uno;M. Fromm;T. Hamakubo;K. Yagi
中科院分区：
文献类型：
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作者：
A. Takahashi;Yumiko Saito;M. Kondoh;K. Matsushita;S. Krug;Hidehiko Suzuki;H. Tsujino;Xiangru Li;H. Aoyama;Koji Matsuhisa;T. Uno;M. Fromm;T. Hamakubo;K. Yagi
claudin （CL）是一个四跨膜蛋白家族，是紧密连接（TJ）的结构和功能成分。由于其在粘膜药物吸收和癌症中的作用，CLs是药物开发的有希望的靶点。然而，由于CL的抗原性低，制备CL蛋白困难，因此CL靶向药物的开发一直被推迟。我们利用产气荚膜梭菌肠毒素（C-CPE）的c端片段和杆状病毒展示系统开发了一种CL结合剂。利用CL-display出芽杆状病毒（BV）从C-CPE突变体显示文库中筛选CL结合物后，我们分离出一个C-CPE突变体m19，它与CL1、CL2、CL4和CL5结合。三维分析表明，m19具有与C-CPE相似的结构骨架。m19和C-CPE的cl结合域的电荷密度不同，表明m19和cl之间可能存在静电相互作用。用m19处理上皮细胞降低了细胞旁完整性，但没有降低跨细胞完整性，m19增强了空肠吸收。因此，我们成功地创建了具有广泛特异性的CL结合剂。这些发现将有助于未来CL靶向药物开发的CL结合剂的制备。
Claudins (CL) are a family of tetra-transmembrane proteins that are the structural and functional components of tight junctions (TJ). CLs are promising targets for drug development because of their role in mucosal drug absorption and cancer. However, CL-targeted drug development has been delayed because CLs have low antigenicity and preparing CL proteins is difficult. We developed a CL binder by using the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE) and a baculoviral display system. After screening CL binders from a C-CPE mutant-displaying library by using CL-displaying budded baculovirus (BV) we isolated a C-CPE mutant called m19, which bound to CL1, CL2, CL4 and CL5. A 3-dimensional analysis showed that m19 has a structural backbone similar to C-CPE. The charge density of the CL-binding domains of m19 and C-CPE differed, suggesting that electrostatic interactions may occur between m19 and CLs. Treatment of epithelial cells with m19 decreased the paracellular but not transcellular integrity, and m19 enhanced jejunal absorption. Thus, we successfully created a CL binder with broad specificity. These findings will contribute to future preparation of CL binders for CL-targeted drug development.