Construction and expression of an anti-GD2,ScFv-IL-2 fusion protein gene.
- Author:
Jianfeng NI
1
;
Jianfei JI
;
An'guo LÜ
;
Ribo HUANG
;
Yutuo WEI
;
Wenfang WU
Author Information
1. Group of Microbiological Engineering, Institute of Applied Ecology, Chinese academy of Sciences, Shenyang 110016, China. nijianfeng2000@126.com
- Publication Type:Journal Article
- MeSH:
Antibodies;
genetics;
metabolism;
Antibodies, Monoclonal;
biosynthesis;
genetics;
Base Sequence;
Cloning, Molecular;
Escherichia coli;
genetics;
metabolism;
Gangliosides;
immunology;
Humans;
Immunoglobulin Fragments;
biosynthesis;
genetics;
immunology;
Interleukin-2;
biosynthesis;
genetics;
Molecular Sequence Data;
Recombinant Fusion Proteins;
biosynthesis;
genetics
- From:
Journal of Biomedical Engineering
2007;24(1):170-175
- CountryChina
- Language:Chinese
-
Abstract:
By combining interleukin2 (IL-2) with a tumor specific antibody, immunotherapy of tumors may become more effective in the future. Anti-GD2 single chain antibody directed to the extracellular domain of GD2 disialoganglioside can result in an antitumor response in some pateins with tumors expressing GD2. In this study, the fusion protein consisting of GD2 single chain antibody (ScFv) and IL-2(Ala125) was constructed. Anti-GD2 ScFv and IL-2 genes were obtained by PCR, then the ScFv-IL-2 gene was constructed by over lap PCR. The gene was inserted into the pMD18-T easy vector. Genes from pMD18-T -vector were inserted into expression vector pSE380. Recombinant expression vector was identified by restriction enzyme-cutting and then was transformed into BL21. SDS-PAGE and Western blot analysis confirmed that the transformed E. Coli BL21 could express ScFv-IL-2 fusion-proteins and the molecular weight is 43 kDa. The fusion protein was purified by affinity chromatograph and Sephacryl S-200HR then was identified through ELISA. The results show that the fusion protein retains the activities of both antigen binding and IL-2.
