The Improvement of Recombinant PirA Antigen Expression in Escherichia coli M15
Summary: Acute Hepatopancreatic Necrosis Disease (AHPND), previously name Early Mortality Syndrome (EMS), is a disease in shrimps which is caused by Vibrio spp. This disease has led to serious shrimp mortality (up to 100%) and caused significant economic losses to shrimp industry. Many studies have shown that AHPND was caused by Vibrio spp. carrying PirA and PirB toxin genes located on the same large plasmid (63-70 kb); they encoded for a binary protein PirAB, which was proved to be the virulence factor of this disease. This research was carried out to evaluate the effect of different environmental conditions during induction on growth of recombinant Escherichia coli M15 and the expression of recombinant PirA antigens. The recombinant E. coli M15 strain carrying the pirA gene derived from our previous research. The gene encoding the pirA antigen was expressed as a fusion protein containing histidine tag at the N-terminal end of the peptide using pQE30 (pQE30-6xHis-PirA) cloned in E. coli M15 cells. Isopropyl β-D-1-thiogalactopyranoside (IPTG) concentration, induction time, and cell density before induction, were selected as three important factors to optimize. The expression levels of 6xHis-PirA fusion protein under the different growth conditions were analyzed. SDS-PAGE analyses was carried out for further confirmation of interest-gene expression. The highest level of 6xHis-pirA fusion protein in recombinant E. coli 15 was determined after induction by 1.00 mM IPTG, for 4 hours after induction, when cell density (OD600nm) reached a value of 1.0 (at 37°C, 200 rpm). From the present outcomes, we conclude that optimizing the three important factors of IPTG concentration, induction time, and cell density before induction can improve 6xHis-pirA fusion protein yield in the E. coli M15 expression system.
Type: SCOPUS listed journals
Author: Nguyễn Văn Khanh, Nguyễn Quang Linh, Nguyễn Thị Bích Phượng, Nguyễn Văn Huệ, Nguyễn Thị Thanh Xuân, Võ Phước Khánh, Trần Quốc Dung
Unit: Laboratory of Protein and Enzyme technology
Journal:Stem Cell
Issue, Number, Pages11, 3, 9-16
ISSN/ISBN:ISSN: 1945-4570 (print); ISSN: 1945-4732 (online)
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Year of publication: 2020
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