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A kind of antibacterial polypeptide hf-18 and its preparation method and application

A HF-18, antibacterial peptide technology, applied in the direction of antibacterial drugs, peptides, peptide sources, etc., can solve the problems of increased clinical use of antibiotics, slow and difficult discovery of antibacterial drugs, etc., to relieve inflammation, and design and preparation methods are simple and convenient. , the effect of improving antibacterial activity

Active Publication Date: 2022-04-26
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, research data show that anti-infective new entity drugs reached a peak in the 1990s and then declined sharply, and the discovery of antibacterial drugs was slow
About 20 million people die from drug-resistant bacterial infections every year in the world, but in recent decades, the abuse of clinical antibiotics has led to the emergence of drug-resistant bacteria worldwide, which has added difficulties to the clinical use of antibiotics

Method used

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  • A kind of antibacterial polypeptide hf-18 and its preparation method and application
  • A kind of antibacterial polypeptide hf-18 and its preparation method and application
  • A kind of antibacterial polypeptide hf-18 and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Preparation of antibacterial polypeptide HF-18:

[0046](1) Using the polypeptide HFIAP-1 as a template, intercept the region where its sequence is conserved and the α-helix is ​​highly concentrated, and intercept from the first position of the N-terminus to reduce the number of polypeptide amino acids from 37 to 18, reducing the cost of synthesis;

[0047] (2) Using the truncated 18 amino acids as a template, replace histidine at position 12 with positively charged lysine and glycine at position 14, which is not conducive to α-helix formation, with hydrophobic phenylalanine , the overall positive charge was increased from 7 to 8, and the hydrophobicity value was increased from 0.207 to 0.244, thereby obtaining the amino acid sequence of the polypeptide HF-18.

[0048] (3) Polypeptide HF-18 synthesized by solid phase synthesis

[0049] The amino acid sequence of the prepared HF-18 is: glycine-phenylalanine-phenylalanine-lysine-lysine-alanine-tryptophan-arginine-lysine-...

Embodiment 2

[0053] Purity determination (HPLC method) and mass spectrometry analysis results of polypeptide HF-18:

[0054] Example 1 Polypeptide HF-18 was synthesized and purified to obtain a finished product, which was identified by high performance liquid chromatography and mass spectrometry.

[0055] Liquid chromatography analysis conditions: C18 chromatographic column (4.6×250mm, 5 μm); mobile phase A is acetonitrile solution containing 0.1% trifluoroacetic acid, and mobile phase B is pure water containing 0.1% trifluoroacetic acid. The detection wavelength is 220nm; the flow rate is 1.0ml / min; the injection volume is 20μl, and gradient elution is performed. See Table 1 for gradient elution conditions.

[0056] Table 2 gradient elution conditions

[0057]

[0058] Depend on figure 1 It can be seen that the purity of the polypeptide HF-18 prepared in Example 1 of the present invention is greater than 98%.

[0059] Depend on figure 2 To determine, the molecular weight of HF-18...

Embodiment 3

[0061] Determination of antibacterial activity of polypeptide HF-18 of the present invention in vitro

[0062] (1) Recovery and activation of strains

[0063] The experimental strains frozen at -20°C were transferred from the glycerol tube to the corresponding agar slant medium (bacteria to nutrient agar slant, fungi to Sabouraud dextrose agar slant). Bacteria were cultured at 37°C for 24 hours, fungi were cultured at 25°C for 48 hours, and placed in a refrigerator at 4°C for later use.

[0064] (2) Preparation of bacteria solution

[0065] Inoculate a little of the bacteria in the slant, transfer to 2ml of the corresponding liquid medium, culture at 37°C or 25°C for 8 hours, and dilute to 10% with liquid medium 5 CFU / ml bacterial suspension, for later use.

[0066] (3) Drug preparation

[0067] The polypeptide HF-18 prepared in Example 1 and HFIAP-1 were weighed and dissolved in physiological saline respectively, and both were prepared into a mother solution of 1024 μg / ml...

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Abstract

The present invention discloses an antibacterial polypeptide HF-18 and its preparation method and application. The amino acid sequence of the antibacterial polypeptide HF-18 is shown in SEQ ID NO:1. The present invention starts from the hagfish polypeptide HFIAP‑1, and obtains a polypeptide of 18 amino acids with a better α-helix degree through the study of the structure-activity relationship, and replaces histidine at the 12th position with lysine and lysine at the 14th position. Glycine was replaced by hydrophobic phenylalanine to obtain polypeptide HF-18 (GFFKKAWRKVKKAFRRVL). The polypeptide HF-18 prepared by the present invention has improved α-helix degree and hydrophobicity, reduced synthesis cost, good antibacterial activity in vivo and in vitro, can be applied to various common infections and stubborn infectious diseases, and can be used as an excellent substitute for existing antibiotics Drugs or Complementary Medicines.

Description

technical field [0001] The invention belongs to the technical field of polypeptide drugs in biochemistry, and in particular relates to an antibacterial polypeptide HF-18 and its preparation method and application. Background technique [0002] Infectious diseases, as a disease with high morbidity and high fatality rate, seriously threaten human health. Antibiotics have always been the first choice for humans to fight bacterial infections. However, research data show that anti-infective new entity drugs reached a peak in the 1990s and then declined sharply, and the discovery of antibacterial drugs was slow. About 20 million people die from drug-resistant bacterial infections every year in the world, but the abuse of clinical antibiotics in recent decades has led to the emergence of drug-resistant bacteria worldwide, which has added difficulties to the clinical use of antibiotics. [0003] Peptides are an important part of the body's innate immune system. They are ubiquitous ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/46A61K38/17A61P31/04
CPCA61P31/04C07K14/461A61K38/00Y02A50/30
Inventor 周长林吴皓旻蒋美玲马菱蔓黄亚杨晓倩
Owner CHINA PHARM UNIV
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