Antisera preparation and epitope mapping of a recombinant protein comprising three peptide fragments of the cystic fibrosis transmembrane conductance regulator

作者：Li Kun; Tang Haiping; Xu Wanxiang; Chen Aijun; Shi Qixian; Sun Zhida; Wang Liyan; Ni Ya^*

来源：Protein Expression and Purification, 2015, 114: 23-29.

DOI：10.1016/j.pep.2015.06.005

• CFTR
• B cell epitope
• Epitope mapping
• Polyclonal antiserum
• Antibody
• Protein expression

Antibodies targeting a single epitope of the cystic fibrosis transmembrane conductance regulator (CFTR) have been reported to influence the validity of immunological analyses; however, autoimmune mechanisms associated with CFTR epitopes are not well understood. In this study, antiserum raised against a multi-epitope recombinant protein composed of three peptide fragments of CFTR (r-CFTR-3P) was prepared and B cell epitope mapping of the protein was carried out using biosynthetic peptides. The r-CFTR-3P gene was cloned into the pSY621 expression plasmid and the protein was expressed in the BL21 strain of Escherichia coli. The rabbit r-CFTR-3P antiserum recognized the native CFTR antigen extracted from human sperm and the GST188 fusion peptides CFTR25-36, CFTR103-117, and CFTR1387-1480 spanning different regions of CFTR. Four novel r-CFTR-3P B cell epitopes were identified: (29)RQRLEL(34), (104)RIIASY(109), (PDN113)-P-111, and (VKLF1450)-V-1447 or CFTR. Other proteins from various species shared sequence homology with the identified epitopes based on NCBI BLAST alignment. This study provides new tools for detecting CFTR protein and insight into the characteristics of minimal B cell epitopes of CFTR and associated immunological mechanisms.

• 出版日期2015-10
• 单位浙江省医学科学院; 上海市计划生育科学研究所