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Low-cost high throughput screening method of antibacterial peptide lead compound

A lead compound, high-throughput technology, applied in the fields of peptide preparation, chemical instruments and methods, library screening, etc., to achieve the effect of simple and feasible process, improved speed and high efficiency

Inactive Publication Date: 2014-02-19
刁有江
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The existing development technology of antibacterial drugs has the following disadvantages: 1) The development of traditional drugs is generally to isolate chemical substances with specific pharmacological effects from nature, and use them as lead compounds for drug research, and then further design, transform and synthesize effective compounds. Functional drugs; and for proteins (polypeptides), they are generally transformed into genetically engineered drugs by means of genetic engineering
Although molecular biology experimental techniques have made great progress, the discovery of drug targets is the bottleneck of development

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0022] 1. Experimental materials

[0023] 1. Strains

[0024] Escherichia coli ( Escherichia coli ) TG1 and the helper phage M13KO7 were preserved in the laboratory of Shandong Agricultural Engineering College.

[0025] 2. Main reagents

[0026] The phage display 10-peptide library was constructed for the laboratory of Shandong Agricultural Engineering College (stock capacity 1.36×10 8 ), the HRP-M13 antibody was purchased from Huamei Company, the 96-well microtiter plate was produced by Nunc Company, and the ELISA reagents were all produced by Huamei Company.

[0027] Preparation of the main solution:

[0028] PBS:

[0029] 1×PBS: 133mmol / l NaCl

[0030] 3mmol / L KCl,

[0031] 8mmol / L Na 2 HPO 4

[0032] 1.5mmol / L KH 2 PO 4

[0033] 1 0×PBS: 80.0gNaCl

[0034] 2.0gKCl

[0035] 11.5gNa 2 HPO 4 ·7H 2 o

[0036] 20.0gKH 2 PO 4

[0037] dissolved in 1 L of H 2 O, adjusted to pH 7.5, and autoclaved.

[0038] PBS-Tween 20 (PBST):

[0039] Add 1ml Tween-20 t...

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PUM

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Abstract

The invention belongs to the technical field of polypeptide drugs, and particularly relates to a low-cost high throughput screening method of an antibacterial peptide lead compound. The method is characterized in that a bacterial cell is taken as a target cell, polypeptide specifically bonded with the surface of the bacterial cell is panned from a phage display random peptide library, and the antibacterial peptide lead compound is screened in a high throughput manner after positive clone, ssDNA (single-stranded deoxyribonucleic acid) sequencing, synthesis of affinity peptides and determination of antibacterial activity; an escherichia coli cell is taken as a target, polypeptide specifically bonded with the surface of the bacterial cell is panned from the phage display random peptide library, and the amino acid sequence of a decapeptide affinity peptide is QKRPRVRLSA. According to the method, polypeptide with specific biological activity, such as an antimicrobial peptide and the like, can be obtained through high throughput screening from random peptides with specific lengths with an affinity screening method in a phage peptide library, the antibacterial peptide lead compound can be rapidly screened with the method, and the development speed of new medicine for the antimicrobial peptide is increased.

Description

technical field [0001] The invention belongs to the technical field of polypeptide drugs in biochemistry, and in particular relates to a low-cost, high-throughput method for screening antimicrobial peptide lead compounds. Background technique [0002] The rapid growth and spread of pathogenic bacteria's resistance to clinical antimicrobial drugs has become one of the most serious problems endangering public health. The emergence of multidrug-resistant Escherichia coli has caused serious difficulties in clinical treatment. The development of safe and effective new antibacterial drugs that are not prone to drug resistance is an important issue before us. [0003] The development of traditional drugs is generally to isolate chemical substances or proteins (polypeptides) with specific pharmacological effects from animals, plants and microorganisms in nature, as the lead compounds for drug research, and then further design, modify and synthesize effective functional drugs ; and...

Claims

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Application Information

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IPC IPC(8): C40B30/06C07K7/06C07K1/14
Inventor 刁有江
Owner 刁有江
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