Preparation method and application of interfering peptide targeting SARS-CoV-2 N protein

A sars-cov-2n and protein technology, applied in the field of preparation of interfering peptides, can solve the problems of limited plasma sources and inability to use on a large scale

Active Publication Date: 2021-07-06
SUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because plasma varies from person to person, it is a complex mixture, an

Method used

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  • Preparation method and application of interfering peptide targeting SARS-CoV-2 N protein
  • Preparation method and application of interfering peptide targeting SARS-CoV-2 N protein
  • Preparation method and application of interfering peptide targeting SARS-CoV-2 N protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Synthesis and detection of interfering peptide drug NIP-V

[0042] The NIP-V amino acid sequence of the interference peptide drug designed by the present invention is IHKNLLIVQDKFPPRRRQRRKKRG, and the targeting sequence is as follows: figure 1 As indicated, D-amino acid was used as a raw material to synthesize at Jill Biochemical (Shanghai) Co., Ltd.

[0043] Such as figure 2 As shown, the synthetic interfering peptide drug NIP-V is identified by the Agilent-6125B liquid mass spectrometry system (Agilent Technologies) as having a molecular weight of 3040.69, and the HPLC adopts an Inertsil ODS-SP liquid chromatography column (Shimadzu, 4.6mm × 250mm) as a stationary phase , using mobile phase A (100% nitrile, 0.1% trifluoroacetic acid) and mobile phase B (100% ultrapure water, 0.1% trifluoroacetic acid) for gradient elution, such as image 3 And shown in Table 1, through HPLC identification, purity is greater than 98%.

[0044] keep time content(%) Pea...

Embodiment 2

[0047] Treatment of ACE2 transgenic mice with the interfering peptide drug NIP-V can significantly reduce the proliferation of SARS-CoV-2 in mice.

[0048] 1 Experimental materials

[0049] ACE2 transgenic mice, Daan gene novel coronavirus (2019-nCoV) nucleic acid detection kit (fluorescent PCR method), SARS-CoV-2, NIP-V interfering peptide medicine prepared in Example 1.

[0050] 2 Experimental methods

[0051] Use sterilized PBS to dissolve the NIP-V interfering peptide drug to a concentration of 1 mg / mL. ACE2 transgenic mice were divided into 4 groups with 8 mice in each group. The first and third groups were injected with 0.5 mL sterilized PBS as a control, and the second and fourth groups were injected with 0.5 mL (0.5 mg) NIP-V drug. One hour later, all four groups of mice were anesthetized and intranasally inoculated with SARS-CoV-2, each mouse was inoculated with approximately 1×105 TCID50 virus. After 16 hours and 24 hours of virus infection, the lung tissues of th...

Embodiment 3

[0059] Treatment of ACE2 transgenic mice with the interfering peptide drug NIP-V can significantly inhibit the lung lesions caused by SARS-CoV-2 infection.

[0060] 1 Experimental materials

[0061] ACE2 transgenic mice, SARS-CoV-2, the NIP-V interfering peptide drug prepared in Example 1. Reagents related to tissue fixation, embedding and HE staining (Shenggong).

[0062] 2 Experimental methods

[0063] Use sterilized PBS to dissolve the NIP-V interfering peptide drug to a concentration of 1 mg / mL. ACE2 transgenic mice were divided into 3 groups, the first group was not infected, the second group was injected with 0.5 mL sterilized PBS, and 1 hour later intranasally inoculated with 1×10 5 TCID50 of SARS-CoV-2, the third group was injected with 0.5 mg of NIP-V drug, and 1 hour later intranasally inoculated 1×10 5 TCID50 of SARS-CoV-2. After 24 hours of virus infection, the mouse lung tissue was taken, placed in 4% paraformaldehyde / PBS for tissue fixation, and paraffin sec...

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Abstract

The invention discloses a preparation method of an interfering peptide targeting SARS-CoV-2N protein. The method comprises the following steps: designing an interfering peptide fragment targeting amino acid located in a dimerization structural domain of the SARS-CoV-2N protein; fusing the interfering peptide fragment with HIV-TAT (Human Immunodeficiency Virus-TAT); modifying the interfering peptide fragment fused with HIV-TAT into a retroisomer to obtain the amino acid sequence of the final interfering peptide NIP-V; and synthesizing the interfering peptide NIP-V by using D-type amino acids as raw materials. The interfering peptide drug NIP-V can interact with the dimerization structural domain of the SARS-CoV-2N protein to inhibit oligomerization of the N protein, so that inhibition of the N protein on innate immunity is relieved, and the purpose of inhibiting replication of the SARS-CoV-2 virus in cells and animal bodies is achieved.

Description

technical field [0001] The invention belongs to the technical field of medicine preparation, and in particular relates to a preparation method and application of an interference peptide targeting SARS-CoV-2 N protein. Background technique [0002] SARS-CoV-2 belongs to the genus of coronavirus (Coronavirus, CoV), its basic structure consists of spike (spike, S) protein, envelope (envelope, E) protein, membrane (membrane, M) protein, nucleocapsid (nucleocapsid) , N) protein and genomic single-stranded RNA composition. The N protein is the core component of the virion. The SARS-CoV-2 N protein has a total length of 419 amino acids and is mainly composed of the N-terminal RNA binding domain, the C-terminal dimerization domain and other sequences connecting the two domains. The full sequence of the SARS-CoV-2 N protein contains multiple conserved positive charge distribution regions that bind to RNA, which binds to the viral genome RNA and packages the RNA into a ribonucleocaps...

Claims

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Application Information

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IPC IPC(8): C07K14/00C07K1/00A61K38/16A61P31/14
CPCC07K14/001A61P31/14A61K38/00A61P11/00C07K14/005C12N2770/20022C12N2770/20033C12N2740/16322C07K19/00C07K2319/10C12N2740/16022C12N2740/16071C12N2770/20032C12N2770/20051C12N2770/20071
Inventor 周芳芳代通王帅张龙
Owner SUZHOU UNIV
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