Method for purifying endogenous human HDAC11 protein inclusion body

By constructing PEGX-6p-1-GST-homo-HDAC11 (designal peptide) recombinant plasmid, Rosetta bacteria induction and inclusion body protein purification, the low protein expression level during the purification of endogenous human HDAC11 protein inclusion body was solved. , the problem of difficult dissolution of inclusion bodies and low regeneration efficiency has achieved efficient protein purification and biological activity verification.

CN119464342APending Publication Date: 2025-02-18CHONGQING MEDICAL UNIVERSITY
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Patent Information

Application Number
CN202411757798.4
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2024-12-03
Publication Date
2025-02-18

AI Technical Summary

Technical Problem

In the prior art, the purification of endogenous human HDAC11 protein inclusion bodies has problems such as low protein expression, difficulty in dissolving the inclusion bodies, and low regeneration efficiency.

Method used

The recombinant plasmid of PEGX-6p-1-GST-homo-HDAC11 (designal peptide) was constructed, Rosetta bacteria induction, inclusion body protein purification, GST-Beads enriched GST-homo-HDAC11 (designal peptide) protein and passed SDS -PAGE and Western Blot verify protein expression and purity.

Benefits of technology

It significantly improves the yield of the target protein, improves the purity of the protein, ensures the biological activity of the protein, and can be used for enzyme activity determination, substrate specific research and the establishment of disease models.

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Abstract

The invention discloses a purification method of an endogenous human HDAC11 protein inclusion body, and relates to the technical field of endogenous human HDAC11 protein inclusion bodies. The purity of the protein inclusion body is improved; the preparation method specifically comprises the following steps: S1, constructing a PEGX-6p-1-GST-homo-HDAC11 (signal peptide removal) vector; s2, carrying out PEGX-6p-1-GST (glutathione S transferase)-homo-HDAC11 (signal peptide removal) Rosetta bacterium induction, and carrying out induction by using the Rosetta bacterium; and S3, purifying the protein of the inclusion body GST-HDAC11 (without signal peptide). The yield of the target protein can be remarkably improved by purifying the inclusion body, the purification process of the inclusion body generally comprises the steps of centrifuging, washing, dissolving and the like, and the steps are beneficial to removing most of cell debris and other impurities and improving the purity of the protein; meanwhile, the purified inclusion body is refolded into the protein with biological activity through denaturation and renaturation methods.
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