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Showing 2 results for Clostridium Botulinum
Dr Seyed Safa Ali Fatemi, Khairollah Yari, Dr Mahmood Tavallaei, Dr Danial Kahrizi,
Volume 16, Issue 2 (8-2011)
Abstract
ABSTRACT
Background and Aim: Botulism is a lethal disease which is caused by the one of the neurotoxins of the 7 types of Clostridium botulinum. The carboxylic domain of the heavy chain of Clostridium botulinum type A neurotoxin (BoNT/A-Hc), is highly capable of activating immune system and is used for production of a vaccine against botulism. The aim of this study was to express and produce soluble form of BoNT/A-Hc in recombinant E. coli.
Materials and Methods: Hc part of BoNT/A was subcloned in pET28a vector and clone was expressed in E. coli BL21 (DE3). Then the best recombinant clones were selected based on three main factors: expression, growth and plasmid stability. Then protein expression was evaluated in LB and M9 media and the protein was purified by resin column chromatography.
Results: In flask culture, under optimal conditions of bacterial culture yielded 52mg of BoNT/A-Hc soluble protein from each liter of culture medium.
Conclusions: According to the results of this study, selection of suitable strains on the basis of important growth indices of the bacteria, expression of recombinant protein and plasmid stability can lead to in increased efficiency of the recombinant protein production.
Key Words: Protein Expresion, E. coli, Clostridium botulinum, Botulism
Conflict of Interest: Nill
Received: Feb 15, 2011 Accepted: May 18, 2011
Mr Hossein Samiei Abianeh, Dr Shahram Nazarian, Dr Mohammad Ebrahim Minaei, Dr Jafar Amani, Mr Amir Sajjad Hojjati Razgi, Dr ,
Volume 28, Issue 3 (8-2023)
Abstract
Background and Aim: Botulism, a syndrome caused by food poisoning, results from use of food contaminated with the botulinum toxin, which is very dangerous and deadly. Botulism is caused by the effects of bacterial toxins on the terminals of the motor nerves. Botulinum neurotoxins are among the most potent toxins. The aim of this study was to investigate expression, purification and, evaluation of the immunogenicity of the recombinant BoNT/B-HcC protein as an immunogen candidate in mice.
Materials and Methods: The C-terminus of the receptor-binding domain of botulinum neurotoxin type B(BoNT/B-HcC) was selected as the antigen for bioinformatics evaluations. The pET17b-BoNT/B-HcC plasmid was transferred to E. coli BL21(DE3) by heat shock. The recombinant protein was purified and analyzed by SDS-PAGE. After verification of the recombinant protein by western blot, immunization of mice was performed. Antibody titers of recombinant proteins were evaluated by indirect ELISA and the results were compared using t-test.
Results: The codon adaptation index (CAI) of the optimized gene was 0.99. The percentage of codons having high-frequency distribution was improved to 78%. Restriction analysis confirmed the 1119 bp construct gene and the recombinant protein with a molecular weight of 43.8 kDa was expressed in the prokaryotic host. The total yield of purified protein was 23 mg of protein per liter of culture. Immunization of mice induced serum antibody response. Statistical analysis showed that the antibody titer was significantly different compared to that of the control sample.
Conclusion: The designed recombinant antigen showed high antigenicity that could be considered as an immunogen against botulinum type B neurotoxin in future studies.
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