Soluble expression method, expression vector and expression engineering bacterium of hirudo nipponia destabilizing enzyme

By constructing fusion proteins with chaperone molecules and TEV restriction sites, soluble expression of the unstable enzyme *Hirudo medicinalis* was achieved in *E. coli*, solving the refolding problem caused by inclusion body expression of the unstable enzyme, improving its bioactivity and preparation efficiency, and promoting its application in biocatalysis and biomedicine.

CN122146670APending Publication Date: 2026-06-05YANCHENG INST OF TECH
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Patent Information

Application Number
CN202610348444.7
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2026-03-20
Publication Date
2026-06-05

AI Technical Summary

Technical Problem

Destabilized enzymes tend to form inclusion bodies in prokaryotic expression systems, leading to cumbersome refolding processes and loss of biological activity, which limits their industrial application in biocatalysis and biomedicine.

Method used

By constructing a fusion protein with a chaperone molecule and a TEV restriction site, soluble expression was achieved in E. coli using the pET-15b vector. Combined with IPTG induction and rTEV protease treatment, the purification steps were simplified, ensuring the enzyme's native conformation and biological activity.

Benefits of technology

This study achieved efficient and soluble expression of unstable enzymes, simplified the purification process, ensured the enzyme's biological activity, reduced preparation costs, and provided technical support for its large-scale application.

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Abstract

The invention provides a soluble expression method, an expression vector and an expression engineering bacterium of hirudo nipponia destabilizing enzyme, and belongs to the technical field of prokaryotic expression. The invention provides a brand-new soluble prokaryotic expression method of the destabilizing enzyme, the soluble high-efficiency expression of the destabilizing enzyme in the supernate is realized by accurately designing a vector construction element, selecting an expression host and the like, the tedious steps of inclusion body renaturation and the folding error risk are avoided, and the method is suitable for large-scale popularization and application. The natural conformation and biological activity of the target enzyme are guaranteed, the subsequent purification process is greatly simplified, the preparation efficiency is improved, a feasible technical scheme is provided for large-scale preparation and practical application of unstable enzymes, and the method has important industrial application value.
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