# Recombinant Detoxified Holotoxin as a Potent Candidate Vaccine Against Botulism

## TL;DR

This study develops a safe and effective recombinant detoxified full-length BoNTA vaccine that shows strong immune responses and protection against botulism in mice.

## Contribution

The novel contribution is the development of a detoxified full-length BoNTA mutant with superior immunogenicity compared to existing approaches.

## Key findings

- The BoNTA/M4 mutant showed the highest safety with a murine lethal dose of 50% (MLD50).
- The M4 protein induced higher antibody levels and better protection against BoNTA challenge than the receptor-binding domain alone.
- The study provides a foundation for a new recombinant detoxified full-length botulinum toxin vaccine.

## Abstract

Vaccination may represent a suitable strategy for preventing botulism. The recombinant expression of toxin functional domains can induce effective immune responses against botulism. This study aims to develop a safe and effective recombinant detoxified full-length BoNTA vaccine. In this study, we engineered and mutated the toxin activity-related sites on the basis of the full-length BoNTA protein and constructed three detoxified full-length toxin mutants. They were recombinant expressed and purified in Escherichia coli, and the BoNTA/M4 mutant was determined to have the highest safety, with a murine lethal dose of 50% (MLD50). The M4 protein was used as the antigen for three immunizations, and the serum titers, neutralizing activity, and BoNTA protective effects of immunized mice were evaluated. The results show that, in comparison to the receptor-binding domain BoNTA/Hc protein, the full-length detoxified mutant M4 protein exhibited superior immunogenicity and could induce higher levels of specific antibodies, and the resulting immune serum could effectively protect mice against higher doses of BoNTA challenge. This study laid the foundation for research on a novel recombinant detoxified full-length botulinum toxin vaccine.