Molecular Cloning and Expression of Novel Fibroblast Growth Factor-2 Conjugated With Immunodominant Domains of Pseudomonas Exotoxin Publisher



Haghighatfard H^{1, 2} ; Samaei NM¹ ; Farazmandfar T¹ ; Hakemi MG³ ; Yamchi A⁴ ; Jadidiniaragh F^{5, 6} ; Yazdani Y^{1, 2} Show Affiliations
Source: Biomedical and Pharmacology Journal Published:2015

Abstract

Angiogenesis is very important in cancer growth and metastasis. Basic fibroblast growth factor (bFGF) as one of the most important angiogenesis factors is an attractive target for cancer vaccine. Due to low immunogenicity, it cannot stimulate an effective immune response. Theoretically, pseudomonas exotoxin (PE) as a potent immunogenic carrier protein when fused to low immunogenic antigens such as bFGF significantly increased immunogenicity of it. In this study, we tried to molecular cloning and expression of bFGF conjugated with immunodominant domains of pseudomonas exotoxin. The coding sequence of fusion protein composed of bFGF linked to PE domains 1b and 2 using EAAAK poly linker. The KDEL sequence was also used in C-terminal coding sequence. It was synthesized and expressed using recombinant DNA technology in the bacterial expression system. Expression of recombinant protein verified using SDS-PAGE and western blot analyses. Finally, it purified using Ni-affinity chromatography. The band close to 37 kDa in SDS-PAGE and western blot analyses was aligned completely to designed sequence. Purified recombinant protein also showed as a clear single band near to 37 kDa.