High-level expression of the Hcc domain of Clostridium botulinum neurotoxin serotype A in Escherichia coli and its immunogenicity as an antigen.
- Author:
Yun-Zhou YU
1
;
Zhi-Wei SUN
;
Shuang WANG
;
Wei-Yuan YU
Author Information
1. Beijing Institute of Biotechnology, Beijing 100071, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Antibodies, Bacterial;
blood;
Bacterial Vaccines;
genetics;
immunology;
Botulinum Toxins, Type A;
biosynthesis;
genetics;
immunology;
Botulism;
immunology;
prevention & control;
Clostridium botulinum type A;
genetics;
immunology;
Escherichia coli;
genetics;
metabolism;
Female;
Lymphocyte Activation;
Mice;
Mice, Inbred BALB C;
Recombinant Proteins;
biosynthesis;
genetics;
immunology;
T-Lymphocytes;
immunology;
Vaccines, DNA;
genetics;
immunology
- From:
Chinese Journal of Biotechnology
2007;23(5):812-817
- CountryChina
- Language:English
-
Abstract:
A completely synthetic gene encoding the He domain of Clostridium botulinum neurotoxin serotype A (AHc, 1287 bp, 429 aa, -50 kD) was constructed with oligonucleotides. After expressed in Escherichia coli, soluble product AHc was gained and verified by SDS-PAGE and Western blot analysis. The expressive level of recombinant AHc in E. coli was very high (36%-53% of soluble total proteins) and the purified yield was more than 30 mg/L by one-step purification. Then, the purified AHc was used to vaccinate Balb/c mice, which developed a strong and specific immune response as expected following administration of AHe protein via the subcutaneous route. Results from BoNT/A neutralization assay showed that the serum from mice vaccinated with AHc contained high titer protective antibody. These results showed that the soluble, stable and high-levelly expressive AHc not only could be produced by the prokaryotic expression system built in our lab, but also owned strong immunogenicity to prepare antitoxin for treatment and as sub-unit candidate vaccine for prophylaxis against botulinum toxin serotype A.
