Kamal Veisi
1,2,3, Safar Farajnia
1,4*, Nosratollah Zarghami
2, Hamid Reza Khoram Khorshid
5, Nasser Samadi
4,2, Shiva Ahdi Khosroshahi
4, Hossein Zarei Jaliani
61 Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
2 Department of Medical Biotechnologies, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran.
3 Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran.
4 Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
5 University of Social Welfare and Rehabilitation Sciences, Tehran, Iran.
6 Department of Medical Genetics, School of Medicine, Shahid Sadoughi University of Medical Sciences, Yazd, Iran.
*Corresponding Author: Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. , Email:
farajnias@tbzmed.ac.ir
Abstract
Purpose: Formation of inclusion bodies is a
considerable obstacle threatening the advantages of E. coli expression
system to serve as the most common and easiest system in recombinant protein
production. To solve this problem, several strategies have been proposed among
which application of molecular chaperones is of remarkable consideration. The
aim of this study was to evaluate the effects of molecular chaperones on
soluble expression of aggregation-prone humanized single chain antibody.
Methods: To increase the solubility of a humanized
single chain antibody (hscFv), different chaperone plasmids including PG-tf2
(GroES- GroEL- tig), ptf16 (tig) and pGro7 (GroES- GroEL) were co-expressed in
BL21 cells containing pET-22b- hscFv construct. The solubility of recombinant
hscFv was analyzed by SDS-PAGE. After purification of soluble hscFv by Ni-NTA
column, the biological activity and cytotoxicity of the recombinant protein
were tested by ELISA and MTT assay, respectively.
Results: SDS-PAGE analysis of the hscFv revealed that
chaperone utility remarkably increased (up to 50%) the solubility of the
protein. ELISA test and MTT assay analyses also confirmed the biological
activity of the gained hscFv in reaction with A431 cells (OD value: 2.6) and
inhibition of their proliferation, respectively.
Conclusion: The
results of this study revealed that co-expression of chaperones with hscFv
leads to remarkable increase in the solubility of the recombinant hscFv, which
could be of great consideration for large scale production of recombinant
single chain antibodies.