Method for improving expression level of botulinum toxin light chain

By constructing a recombinant expression system of the wild-type lysis tag NT11 and its mutant mut7 in Escherichia coli and combining it with the overexpression of the molecular chaperone htpG, the problem of efficient expression of botulinum toxin light chain in host bacteria was solved, realizing an efficient and simplified production process and high-activity expression.

CN121717869APending Publication Date: 2026-03-24BEIJING UNIV OF CHEM TECH
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Patent Information

Application Number
CN202511733628.7
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2025-11-24
Publication Date
2026-03-24

AI Technical Summary

Technical Problem

In existing technologies, type A botulinum toxin light chains are difficult to express efficiently in host bacteria, and the use of traditional lysis-promoting tags requires subsequent enzymatic digestion, which increases process costs and complexity.

Method used

Using the wild-type fusion tag NT11 and its mutants, such as mut7, a recombinant expression system was constructed in E. coli through genetic engineering. By combining the overexpression of the molecular chaperone htpG, the expression vector and fusion tag were optimized to achieve efficient expression without enzyme digestion.

Benefits of technology

The expression level of botulinum toxin light chain was increased to 647 mg/L, which is 20-43% higher than that of traditional methods, while maintaining high activity and simplifying the production process.

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Abstract

The invention provides a method for improving the expression level of a meat toxin light chain. The method comprises the following steps: connecting a dissolution-promoting tag to an N end or a C end of meat toxin light chain protein to form soluble meat toxin light chain recombinant protein; the dissolution-promoting tag is a wild dissolution-promoting tag NT11 or a mutant protein mut7 of the wild dissolution-promoting tag NT11, enzyme digestion removal is not needed, the activity of a meat toxin light chain is hardly influenced and interfered, and the fusion protein can keep 93% of substrate enzyme digestion activity. The method further comprises overexpressing a molecular chaperone htpG in the genome of the host bacterium. According to the invention, a BoNT / A-LC high-efficiency expression system is constructed through multi-strategy optimization, so that the yield of BoNT / A-LC is increased by nearly 20% compared with that of wild type NT11-Lc, the yield reaches 647 mg / L through overexpression of a molecular chaperone htpG, and after combinatorial optimization, the yield is 8.5 times that of only expressed BoNT / A Lc, 1.79 times that of NT11-Lc and 1.43 times that of Mut7-Lc.
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