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Antigenic epitope displaying method based on single domain antibody

A single-domain antibody and antigen epitope technology, applied in the biological field, can solve the problems of poor epitope peptide immunogenicity, high cost, complex epitope peptide immunogen technology, etc., and achieve the effect of avoiding technical complexity

Active Publication Date: 2014-11-26
BEIJING VICNOVO SCI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The purpose of the present invention is to establish a new antigenic epitope display method to realize the effective display of antigenic epitopes, to solve the poor immunogenicity of the current chemically synthesized epitope peptides, and to prepare epitope peptide immunogens by chemical coupling. complex, costly problem

Method used

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  • Antigenic epitope displaying method based on single domain antibody
  • Antigenic epitope displaying method based on single domain antibody
  • Antigenic epitope displaying method based on single domain antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Example 1 Single-domain antibody skeleton display based on camel heavy chain antibody (His) 6 gauge

[0069] Enter the PDB database (http: / / www.pdb.org / pdb / home / home.do), enter "camel antibody" in the search box, and find a camel antibody compounded with lysozyme from the structural data in the search box Structural data (1BZQ) of the object, from which the amino acid sequence of a camelid single domain antibody is shown in SEQ ID NO: 1 (as shown in figure 1 Shown, wherein the double dashed part is the CDR3 region).

[0070] Will figure 1 The amino acid sequence of the CDR3 region shown in is replaced by the "HHHHHH" epitope (His) 6 Epitope, the sequence after replacement is shown in SEQ ID NO: 2 (see attached figure 2 ).

[0071] Then, according to the codon usage bias of Escherichia coli, the sequence shown in the above SEQ ID NO: 2 was converted into its coding gene sequence (http: / / www.bioinformatics.org / sms2 / rev_trans.html), and at the same time, the upstream...

Embodiment 2

[0090] Example 2 Human single domain antibody backbone displaying c-myc epitope and its application

[0091] Obtain a human antibody heavy chain variable region amino acid sequence from the GenBank database and literature (J Exp Med 2011, 208(1): 181-193), as shown in SEQ ID NO: 4, and use this sequence as a single domain antibody base sequence. (if attached Figure 5 Shown, wherein the double underlined part is the framework region 2 (FR2); the single underlined part is the CDR3 region).

[0092] will attach Figure 5 The 9th, 10th and 12th amino acids in FR2 of the single domain antibody sequence shown in are replaced with glutamic acid (E), arginine (R) and glycine (G) respectively, and at the same time, their The CDR3 region was replaced with the c-myc epitope sequence, namely "EQKLISEEDL", and the c-myc epitope display protein was constructed as shown in SEQ ID NO: 5, (see attached Figure 6 shown).

[0093] Then, convert the above sequence into its coding gene seque...

Embodiment 3

[0113] Example 3 Display of Human Vascular Endothelial Growth Factor Epitope Using the Single-Domain Antibody Skeleton Based on Sharklein Heavy Chain Antibody

[0114] From the literature (Molecular Immunology 2007; 44:656-665), a shark single domain antibody was found, the amino acid sequence of which is shown in SEQ ID NO: 7, (see Figure 9 As shown, the underlined part is the CDR3 region).

[0115] Will Figure 9 The amino acid sequence of the CDR3 region shown in is replaced by human vascular endothelial growth factor (VEGF) antigen epitope "YPDEIEYIFKP" epitope, and the sequence after replacement is shown in SEQ ID NO: 8, (see appendix Figure 10 , where the underlined part is the epitope of human vascular endothelial growth factor).

[0116] Then, according to the codon usage bias of Escherichia coli, the sequence shown in the above SEQ ID NO: X was converted into its coding gene sequence (http: / / www.bioinformatics.org / sms2 / rev_trans.html), and at the same time, the up...

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Abstract

Disclosed is an epitope display method based on a single domain antibody. The present invention utilizes a CDR3 area of a single domain antibody to display an exogenous epitope. The skeleton protein of such a single domain antibody, on the surface of which the exogenous epitope is displayed, can be used to prepare an antibody for epitope small peptide, and can also be used as a substitute antigen for an exogenous antigen to immunize animals and prepare an antibody for an exogenous antigen. In addition, the epitope display method based on the single domain antibody skeleton can also be used for preparing epitope vaccine.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method and application of using a single domain antibody to display an exogenous antigen epitope. Background technique [0002] Epitope is a part of protein antigen and is the basis of protein antigenicity, and the specificity of antibody is directed against epitope. Since the epitope is generally only composed of a few to a dozen amino acids, the synthesized individual epitope peptide generally has no or only very weak immunogenicity due to its small molecule. In order to enhance the immunogenicity of antigenic epitope peptides, epitope peptides are generally coupled to carrier proteins such as bovine serum albumin, keyhole limpet hemocyanin, tetanus toxoid, etc., or to lipids with adjuvant activity Molecules are covalently linked to form lipopeptides, and antigenic epitope peptides can also be linked to branched polylysine backbones to form a macromolecule with a uni...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00C07K1/107C12N15/62C12N15/10C12N15/70
CPCC07K2317/569C07K16/00C12N15/1044
Inventor 高建恩罗时伟钱军
Owner BEIJING VICNOVO SCI TECH
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