• Original article (Basic research) • Previous Articles     Next Articles

Study on the human scaffold protein FN3 for displaying NT-proBNP epitopes

LI Meng-yang1, YANG Chun-guang2, SUN Shen-xia2, XU Wei-ye2, WANG Xuan-Yu2, HU Xue-jun2, DING Ning2 , QU Peng1   

  1. 1. Department of Anatomy, Medical College, 2. Medical Research Centre, Da Lian University, Dalian116622, China
  • Online:2016-11-28 Published:2016-11-29
  • Supported by:

    National Natural Science Foundation of China, 31370937


Objective · To display NT-proBNP epitopes using human scaffold protein FN3 and to efficiently express a stable recombinant protein with active NT-proBNP epitopes via E.coli expression system. Methods · The FG loop sequence of scaffold protein FN3 was replaced by NTproBNP epitope 12-21 and 13-20 sequences at amino acid residue, respectively, and was cloned into the E.coli expression vector pET28(a)+. The recombinant FN3 proteins displaying NT-proBNP epitopes were expressed in E.coli BL21(DE3) plySs and were purified. The antigen activity was detected and physiochemical properties were analyzed. Results · The expression vector for recombinant protein FN3 displaying NT-proBNP epitope 12-21 and 13-20 sequences at amino acid residue was successfully constructed. The E.coli strain with efficient expression of scaffold protein FN3 displaying NT-proBNP epitopes was obtained. Highly purified recombinant protein with antigen immune activity against NT-proBNP was obtained with nickel column purification and was verified with Western blotting and ELISA. Differential thermal scanning method showed that the recombinant protein maintained the stability of scaffold protein FN3. Plasma stability test indicated that the recombinant protein was a highly stable substitute for antigen protein with the same effect and could significantly extend the degradation cycle (serous half-life). Conclusion · NT-proBNP was displayed successfully via scaffold protein FN3. A substitute for antigen protein of NT-proBNP with the same antigenicity can be simply, rapidly, and efficiently prepared with E.coli expression system.

Key words: FN3, NTproBNP, antigenic epitope