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Novel coronavirus (SARS-COV-2) spike protein binding molecule and applications thereof

A SARS-COV-2 and spike protein technology, applied in the field of medicine and biology, can solve the problems of being difficult to become a special drug for SARS-COV-2, limited therapeutic effect, etc., and achieve good specificity, inhibition of infection and amplification Effect

Active Publication Date: 2020-09-11
SHENZHEN IMMUNOTHERAPY BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the problem that the existing antiviral drugs have no specificity to the novel coronavirus, the therapeutic effect is limited, and it is difficult to become a specific drug for SARS-COV-2, the present invention provides a novel coronavirus (SARS-COV-2) Spike protein binding molecules and applications thereof

Method used

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  • Novel coronavirus (SARS-COV-2) spike protein binding molecule and applications thereof
  • Novel coronavirus (SARS-COV-2) spike protein binding molecule and applications thereof
  • Novel coronavirus (SARS-COV-2) spike protein binding molecule and applications thereof

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Experimental program
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Effect test

Embodiment 1

[0095] Screening of single domain antibodies against SARS-COV-2-Spike protein

[0096] 1.1 Library construction

[0097] 1.1.1 Immunity

[0098] The alpaca was immunized with the Spike-RBD protein of the new coronavirus, and immunized at the 1st, 2nd, 4th, and 6th week, respectively, for a total of 4 immunizations, and each immunization dose was 200ug.

[0099] 1.1.2 Extraction of total RNA

[0100] Take 50ml of alpaca peripheral blood after immunization in the sixth week, separate lymphocytes, extract total RNA of lymphocytes with Trizol, and use UV spectrophotometer to detect the extracted RNA results: OD260 / 280=1.99, OD260 / 230=1.43 , indicating that the extracted RNA was not significantly degraded, and the purity was good; the total RNA concentration was 809.3ng / μL. The extracted total RNA was used for agarose gel electrophoresis, and the results were as follows: figure 1 As shown, two bands of 28S and 18S can be seen.

[0101] 1.1.3 RNA reverse transcription

[0102]...

Embodiment 2

[0151] 1.1 Preparation of Fc fusion protein of SARS-COV-2-Spike protein single domain antibody

[0152] According to the amino acid sequence of the constant region of human immunoglobulin (IgG1) on the protein database Uniprot, the amino acid sequence of the human IgG1-Fc region (SEQ ID NO: 109) was obtained. Obtain the nucleic acid fragment (nucleic acid sequence such as SEQ ID NO: 137) encoding human IgG1-Fc from human PBMC total RNA by reverse transcription PCR, and then obtain the fusion protein of SARS-COV-2-Spike protein single domain antibody and Fc by overlapping PCR The encoding nucleic acid fragment was recombined into the vector pCDNA4 (Invitrogen, CatV86220).

[0153] The successfully constructed pCDNA4 plasmid containing the nucleic acid fragment of the fusion protein of SARS-COV-2-Spike protein single domain antibody and Fc was transfected into HEK293 cells for expression. Specifically, the recombinant expression plasmid was diluted with Freestyle293 medium and ...

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Abstract

The invention relates to the technical field of medical biology, and specifically discloses a novel coronavirus (SARS-COV-2) spike protein binding molecule and applications thereof. The binding molecule can specifically bind the spike protein of SARS-COV-2 and includes at least one immunoglobulin single variable structural domain. The provided binding molecule can specifically bind the SARS-COV-2-Spike protein and effectively block the binding of the SARS-COV-2-Spike protein and human cell ACE2 receptors, so that the infection process of the SARS-COV-2 on cells can be further blocked, and theinfection and amplification of the SARS-COV-2 can be inhibited.

Description

technical field [0001] The invention relates to the field of medical biotechnology, in particular to a novel coronavirus (SARS-COV-2) spike protein binding molecule and an application thereof. Background technique [0002] Novel coronavirus pneumonia (COVID-19) has infected more than 4 million people worldwide, and the number of infected people is still increasing rapidly. There is currently no specific and effective treatment for COVID-19 clinically. Although the epidemic situation in our country has been fully controlled, the epidemic situation abroad has broken out and is still growing rapidly. In addition, more and more studies have shown that the new coronavirus (SARS-COV-2) infection may have a chronic carrier state; some discharged Fuyang patients also suggest that the virus may exist in the human body for a long time. The key factors such as the mechanism and time of long-term carry are still unclear, and it is crucial to prevent the resurgence of SARS-COV-2 in the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10C12N15/13C12P21/02A61K39/395A61K47/68A61P31/14G01N33/569
CPCC07K16/10A61K47/68A61P31/14G01N33/56983C07K2317/569C07K2317/565C07K2317/76C07K2317/92C07K2317/94A61K2039/505G01N2469/20G01N2333/165Y02A50/30C07K2317/52C07K2317/22G01N2469/10G01N27/447G01N2030/8831G01N30/88C07K16/1003C12N15/63
Inventor 张军方
Owner SHENZHEN IMMUNOTHERAPY BIOTECH CO LTD
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