Leading stabilizing element for enhancing activity and expression of exogenous protein in bacteria

A technology for exogenous proteins and stabilizing elements, applied in the biological field, can solve the problems of lack of activity, harmful effects of heterologous proteins, and protein instability, and achieve the effects of enhancing expression and activity, and improving protein yield and protein activity.

Active Publication Date: 2021-10-01
SHANXI UNIV
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Problems solved by technology

Target protein cannot be detected or is detected by sensitive techniques such as Western blot
At very low levels (less than micrograms per liter of culture), the problem is usually either that th...

Method used

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  • Leading stabilizing element for enhancing activity and expression of exogenous protein in bacteria
  • Leading stabilizing element for enhancing activity and expression of exogenous protein in bacteria
  • Leading stabilizing element for enhancing activity and expression of exogenous protein in bacteria

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Embodiment

[0027] A leading stable element that enhances the activity and expression of exogenous proteins in bacteria is a 208-amino acid residue fragment of the N-terminal of the Salmonella enterica subsp. Shown in SEQ.ID.NO:1, the amino acid sequence is shown in SEQ.ID.NO:2.

[0028] Identification of lead stabilizing elements that enhance the activity and expression of foreign proteins in bacteria:

[0029] 1. Cloning the minimal promoter (SEQ.ID.NO: 3) of the Salmonella enterica subsp. enterica Chlamydia typhimurium strain RM13672 SspH1 and the coding sequence of 208 amino acid residues at the N-terminal of the SspH1 protein (a total of 889 nucleotide sequences, see figure 1 shown);

[0030] According to the SspH1 gene coding region design primer pair pSspH1-265-F / pSspH1-R2 in the chromosome sequence of Salmonella enterica subsp. pSspH1-R2, bacterial genomic DNA and high-fidelity Tag DNA polymerase to establish polymerase chain reaction (PCR), the reaction conditions are 95°C for ...

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Abstract

The invention belongs to the technical field of biology, and discloses a leading stabilizing element for enhancing activity and expression of exogenous protein in bacteria. The leading stabilizing element is 208 amino acid residue segments at the N end of the salmonella enteric subspecies chlamydia typhimurium strain RM13672SspH1 protein, the nucleotide sequence is shown in SEQ.ID. NO: 1, and the amino acid sequence is shown in SEQ.ID. NO: 2. the leading stabilizing element for enhancing activity and expression of the exogenous protein in the bacteria can be used as a tool for improving the protein yield and the protein activity in the industry or scientific research, and is beneficial to scientific research, biological medicine industrial production and preparation of anti-infectious disease vaccines. Besides, glutathione S transferase (GST) is a protein containing 211 amino acids, is the most common stable polypeptide expressed by the protein, but has great effect difference on different target proteins. The identified SsppH1 leading stabilizing element can provide a new choice so as to obtain ideal expression of active protein.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a leading stabilizing element for enhancing the activity and expression of foreign protein in bacteria. Background technique [0002] In scientific research and biomedical industry production and anti-infectious disease vaccine preparation, it is often necessary to express or purify biologically active bacteria in prokaryotic bacteria (including but not limited to Escherichia coli, Salmonella, Shigella, Mycobacterium tuberculosis, etc.) protein. [0003] However, the expression of foreign proteins in bacteria often encounters the following problems: [0004] (1) No output or low output. The protein of interest cannot be detected or is detected by sensitive techniques such as Western blot. At very low levels (less than micrograms per liter of culture), the problem is usually either that the heterologous protein is acting deleteriously or that the protein itself is not stable. [0...

Claims

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Application Information

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IPC IPC(8): C07K14/255C12N15/31C12N15/70C12N15/66C12Q1/66
CPCC07K14/255C12N15/70C12N15/66C12Q1/66Y02A50/30
Inventor 邢力潘元晴吴长新
Owner SHANXI UNIV
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