Abrol S, Sampath A, Arora K, Chaudhary V K
Department of Biochemistry, University of Delhi South Campus, New Delhi, India.
Indian J Biochem Biophys. 1994 Aug;31(4):302-9.
The envelope glycoprotein, gp120, on the surface of HIV interacts with the human CD4 molecule and thus helps the virus in gaining entry into the T-helper cells. To display the gp120 binding domains of human CD4 on the surface of the bacteriophage M13, two types of vectors have been constructed. In these, the first 176 amino acids of the human CD4 have been fused with the minor coat protein, gIIIp, of M13 bacteriophage for surface display. The Western blot analysis revealed that using the phage based vector, M13CD41923, all the copies of gIIIP (3-5 per virion) were present as fusion protein indicating multivalent display. In the phagemid based vector, phage particles were produced only upon infection of the cells carrying pVCCD43426, with the helper phage, M13KO7. Thus these phage particles carried both, the fusion protein as well as the unfused gIIIp, as shown by Western blot analysis. The presence of large amount of unfused gIIIp ensured that the phage particles did not display more than one fusion protein per phage particle, thus leading to monovalent display. Phage particles produced by both vectors could be captured on immobilized gp120, thereby showing that the displayed CD4 domains were functional.
人类免疫缺陷病毒（HIV）表面的包膜糖蛋白gp120与人类CD4分子相互作用，从而帮助病毒进入辅助性T细胞。为了在噬菌体M13表面展示人类CD4的gp120结合结构域，构建了两种类型的载体。在这些载体中，人类CD4的前176个氨基酸与M13噬菌体的次要外壳蛋白gIIIp融合，用于表面展示。蛋白质印迹分析表明，使用基于噬菌体的载体M13CD41923，所有的gIIIP拷贝（每个病毒粒子3 - 5个）都以融合蛋白的形式存在，表明是多价展示。在基于噬菌粒的载体中，只有在携带pVCCD43426的细胞被辅助噬菌体M13KO7感染后，才会产生噬菌体颗粒。因此，如蛋白质印迹分析所示，这些噬菌体颗粒同时携带融合蛋白和未融合的gIIIp。大量未融合的gIIIp的存在确保每个噬菌体颗粒不会展示超过一个融合蛋白，从而导致单价展示。两种载体产生的噬菌体颗粒都可以被固定化的gp120捕获，从而表明展示的CD4结构域具有功能。
