【摘要】 本实验以pCANTAB5E噬菌粒为载体,成功构建了较高容量的噬菌体展示随机十肽库,并将其应用于抗原模拟表位的淘选和鉴定。将一种特异识别对虾白斑综合症病毒(Whitespotsyndromevirus,WSSV)的单链抗体A1对十肽库和十五肽库分别进行淘选,结果得到一系列能与单链抗体A1特异性结合的阳性克隆。将这些阳性克隆所编码的多肽氨基酸序列与已知的单链抗体A1的抗原WSSV388片段氨基酸序列做比对,发现多数阳性多肽序列都与WSSV388片段序列的C端一处K????R??R?QS的氨基酸片段相似,由此推论单链抗体A1的模拟抗原表位可能是由该不连续氨基酸片段所构成的构象表位,而非线性表位。研究结果表明,噬菌体展示随机肽库技术是一种用于研究抗原表位结构的有效方法,有助于进一步探讨WSSV的结构蛋白的构象及功能,以及相应单链抗体与细胞受体相互作用的机理。更多还原

【Abstract】 A phage display random decapeptide library with high titer was constructed using pCANTAB 5 E as vector and used to select and identify antigen epitopes. ScFv A1, which specifically binds the shrimp’s White spot syndrome virus (WSSV), was used to pan against the decapeptide library and a random 15mer library, respectively. A series of positive clones that bind specifically to scFv A1 were obtained. The amino acid sequences of the positive peptides were compared with WSSV388 fragment, which is the antigen of scFv A1. We found that most of the sequences of the positive peptides were similar to a fragment,KRRQ, located on C-terminal of WSSV388 fragment. We deduced that the mimic antigen epitopes of scFv A1 were conformational epitopes structured by the discontinuous amino acid fragment, but not by a linear amino acid fragment. These results show that phage display peptide library technology is a powerful method for analyzing the structure of antigen epitopes, which is helpful to further explore conformation and function of the structural proteins of WSSV, and the mechanisms of interactions of the virus with the antiviral antibodies and host cells.更多还原