construction of a synthetically engineered nirb promoter for expression of recombinant protein in escherichia coli
نویسندگان
چکیده
background anaerobic-inducible promoters are alternatives of chemical-inducible promoters for expression of recombinant proteins especially in conditions where chemical induction is not possible or anaerobic conditions are preferable. the nirb promoter is the promoter of the first gene of nir operon in escherichia coli, which encodes nadh-dependent nitrite reductase. this promoter is naturally induced under anaerobic conditions and upregulated by nitrite and nitrate. objectives the current study was carried out to construct a synthetic nirb promoter that does not respond to chemical inducers (nitrite or nitrate), but instead responds to anaerobic induction. for this purpose, a new plasmid was constructed (pfsnirb78-23ltb), which contains a synthetic nirb promoter. the activity of this plasmid was evaluated in e. coli under both aerobic and anaerobic conditions and in response to chemical inducers, nitrite and nitrate. materials and methods a synthetic nirb promoter was firstly cloned into a pkk223 derivative plasmid and then the heat labile toxin b subunit gene (ltb) of entrotoxigenic e. coli was cloned under the control of this promoter. the inducibility of this plasmid in e. coli was measured under anaerobic conditions in the presence or absence of nitrite or nitrate by ganglioside gm1 elisa. results our data showed that this promoter is strongly induced under anaerobic conditions while it showed much lower activity (11%) under aerobic conditions. in contrast to the native promoter, this promoter was not induced by chemical inducers, nitrite or nitrate. conclusions this study showed that the recombinant protein produced under the control of synthetic nirb promoter has critical characteristics such as pentamer formation, receptor recognition ability and conservation of antigenic epitopes. in addition, the data showed anaerobiosis and chemical inducers had no adverse effects on recombinant proteins. based on the results, this synthetic promoter is suitable for use in live delivery vaccines or drug systems and for production of recombinant proteins especially oxygen sensitive proteins.
منابع مشابه
Construction of a Synthetically Engineered nirB Promoter for Expression of Recombinant Protein in Escherichia coli
BACKGROUND Anaerobic-inducible promoters are alternatives of chemical-inducible promoters for expression of recombinant proteins especially in conditions where chemical induction is not possible or anaerobic conditions are preferable. The nirB promoter is the promoter of the first gene of nir operon in Escherichia coli, which encodes NADH-dependent nitrite reductase. This promoter is naturally ...
متن کاملCONSTRUCTION OF RECOMBINANT PLASMIDS FOR PERIPLASMIC EXPRESSION OF HUMAN GROWTH HORMONE IN ESCHERICHIA COLI UNDER T7 AND LAC PROMOTERS
In order to study the periplasmic expression of human growth hormone (hGH) in Escherichia coli, the related cDNA was inserted in two expression plasmids carrying pelB signal peptide, one with lac bacterial promoter and the other with a bacteriophage T7-based promoter. The recombinant plasmids were moved to TG1 and BL21 strains of E. coli, respectively. To induce the expression systems, IPTG and...
متن کاملEnhanced Expression of Recombinant Activin A in Escherichia coli by Optimization of Induction Parameters
Activin A is a member of the transforming growth factor β super family. Because of its extensive clinical usages, its recombinant production is beneficial. In this study, activin A was expressed in E. coli using the pET 21a expression vector. The optimization of the activin A production in E. coli was done by using the response surface methodology (RSM). At this stage, the effect of IPTG and la...
متن کاملConstruction of New Genetic Tools as Alternatives for Protein Overexpression in Escherichia coli and Pseudomonas aeruginosa
Background: Pseudomonas protein expression in E. coli is known to be a setback due to signifi cant genetic variation and absence of several genetic elements in E. coli for regulation and activation of Pseudomonas proteins. Modifi cations in promoter/repressor system and shuttle plasmid maintenance have made the expression of stable and active Pseudomonas protein possible in bot...
متن کاملIsolation and expression of recombinant viral protein (VP2) from Iranian isolates of Infectious Pancreatic Necrosis Virus (IPNV) in Escherichia coli
Infectious Pancreatic Necrosis Virus (IPNV) is a member of the family Birnaviridae that has been linked to high mortalities in salmonids. Bacterial based systems as live vectors for the delivery of heterologous antigens offer a number of advantages as vaccination strategies. VP2 is a structural viral protein of IPNV with immunogenicity effects. In this study IPNV was isolated from diseased fry ...
متن کاملمنابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
jundishapur journal of microbiologyجلد ۷، شماره ۷، صفحات ۰-۰
میزبانی شده توسط پلتفرم ابری doprax.com
copyright © 2015-2023