Glucose regulated protein 78 (GRP78) is a typical endoplasmic reticulum (ER) luminal chaperone having a main role in the activation of the unfolded protein response (UPR). Due to hypoxia and nutrient deprivation in the tumor microenvironment, expression of GRP78 in these cells becomes higher than the native cells which makes it a suitable candidate for cancer targeting. Suppression of survival signals by antibody production against C-terminal domain of GR78 (CGRP) can induce apoptosis of cancer cells. The aim of this study was in silico analysis, recombinant production and characterization of CGRP in E.coli. Structural prediction of CGRP by bioinformatics tools was done and the construct containing optimized se... More
Glucose regulated protein 78 (GRP78) is a typical endoplasmic reticulum (ER) luminal chaperone having a main role in the activation of the unfolded protein response (UPR). Due to hypoxia and nutrient deprivation in the tumor microenvironment, expression of GRP78 in these cells becomes higher than the native cells which makes it a suitable candidate for cancer targeting. Suppression of survival signals by antibody production against C-terminal domain of GR78 (CGRP) can induce apoptosis of cancer cells. The aim of this study was in silico analysis, recombinant production and characterization of CGRP in E.coli. Structural prediction of CGRP by bioinformatics tools was done and the construct containing optimized sequence was transferred to E.coli T7 shuffle. Expression was induced by IPTG and recombinant protein was purified by Ni- NTA agarose resin. The content of secondary structures was obtained by Circular Dichroism spectrum. CGRP immunogenicity was evaluated from the immunized mice sera . SDS-PAGE analysis showed CGRP expression in E.coli. Circular Dichroism spectrum also confirmed prediction of structures by bioinformatics tools. The enzyme-linked immunasorbent assay (ELISA) using sera from immunized mice revealed CGRP as a good immunogen. The results obtained in this study showed that the structure of truncated CGRP, is very similar to its structure in the whole protein context. This protein can be used in cancer researches. This article is protected by copyright. All rights reserved.
