title: Enhanced expression of soluble recombinant tetanus neurotoxin Hc in Escherichia coli as a tetanus vaccine candidate.
abstract: The expression of the carboxyl fragment of the heavy chain of tetanus neurotoxin (TeNT-Hc) in Escherichia coli has been hampered by the unusually high AT content and the presence of rarely used codons by Clostridium. The gene encoding TeNT-Hc was optimized for E. coli by replacing rare codons and decreasing the AT pairs from 72.57% to 52.47%. The reconstructed gene was expressed in E. coli BL21(DE3) and resulted in a soluble product which was about 46% of the total bacterial protein. TeNT-Hc produced in a 42 L fermentor was purified to >95% at 87 g/kg of cell paste (approximately 333 mg/L). BALB/c mice vaccinated with three bi-weekly doses of TeNT-Hc with Freund's adjuvant were fully protected against an intraperitoneally challenge of 2 × 10(3) 50% lethal doses (LD(50)s) of tetanus neurotoxin. NIH mice vaccinated with TeNT-Hc adsorbed to aluminum hydroxide gel adjuvant demonstrated a potency of 168 IU/mL, which was 2 times higher than the national standard for tetanus vaccines. These results suggest that TeNT-Hc may be a promising second-generation vaccine candidate for clinical use against tetanus neurotoxin.
authors.affiliation: State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, 20 Dongdajie Street, Fengtai, Beijing 100071, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Yu
authors.forename: Lihua
authors.initials: L
authors.lastname: Hou
authors.forename: Changming
authors.initials: C
authors.lastname: Yu
authors.forename: Shuling
authors.initials: S
authors.lastname: Liu
authors.forename: Jun
authors.initials: J
authors.lastname: Ren
authors.forename: Ting
authors.initials: T
authors.lastname: Fang
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Zhang
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
mesh_terms: D000276:Adjuvants, Immunologic; D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D005732:Gangliosides / Q000276:immunology; D023061:Gene Order; D008297:Male; D008666:Metalloendopeptidases / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D012995:Solubility; D013742:Tetanus / Q000517:prevention & control; D013744:Tetanus Toxin / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D013745:Tetanus Toxoid / Q000276:immunology*; D014611:Vaccination; D022223:Vaccines, Subunit / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D057134:Antibodies, Neutralizing; D005732:Gangliosides; D011994:Recombinant Proteins; D013744:Tetanus Toxin; D013745:Tetanus Toxoid; D022223:Vaccines, Subunit; C035979:tetanospasmin; C016481:trisialoganglioside GT1; D008666:Metalloendopeptidases