Antibacterial peptide P104 and lyase LysP53 with broad-spectrum splitting activity, and application of antibacterial peptide P104 and lyase LysP53

A technology of P104 and lyase, which is applied in the direction of lyase, medical preparations containing active ingredients, applications, etc., can solve the problems of less research on phage lyase, and achieve high enzyme activity, good cleavage effect, and good application prospects Effect

Active Publication Date: 2021-10-26
WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the application of phage lyase is mainly to lyse Gram-positive bacteria, such as Staphylococcus aureus, Listeria and Enterococcus, etc. There are few studies on phage lyase with lytic activity on Gram-negative bacteria

Method used

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  • Antibacterial peptide P104 and lyase LysP53 with broad-spectrum splitting activity, and application of antibacterial peptide P104 and lyase LysP53
  • Antibacterial peptide P104 and lyase LysP53 with broad-spectrum splitting activity, and application of antibacterial peptide P104 and lyase LysP53
  • Antibacterial peptide P104 and lyase LysP53 with broad-spectrum splitting activity, and application of antibacterial peptide P104 and lyase LysP53

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] The synthesis of embodiment 1 antimicrobial peptide P104

[0043] After a large number of experimental analysis by the inventor, the inventor found a section of antimicrobial peptide P104 from the phage P53 genome, the amino acid sequence of the antibacterial peptide P104 is shown in SEQ ID NO.1; the gene sequence of the antibacterial peptide P104 is shown in SEQ ID NO. 3. The antimicrobial peptide P104 was synthesized by Shanghai Qiangyao Biotechnology Co., Ltd. The purity of the synthesized antimicrobial peptide P104 was 98.58%, and the molecular weight was 3885.57. The purity of the antimicrobial peptide P104 was determined by high performance liquid chromatography. The chromatographic purity of the antimicrobial peptide P104 picture see figure 1, and the analysis results are shown in Table 1 below,

[0044] Table 1 Antimicrobial Peptide P104 Liquid Chromatography Detection Component List

[0045] retention time (min) Peak area purity(%) 12.085 ...

Embodiment 2

[0050] Example 2 Expression and purification of lyase LysP53

[0051] After a large number of experimental analysis by the inventor, the inventor found a lyase LysP53 from the phage P53 genome, the amino acid sequence of the lyase LysP53 is shown in SEQ ID NO.2; the gene sequence of the lyase LysP53 is shown in SEQ ID NO .4 shown.

[0052] 2.1 Construction of recombinant expression vector

[0053] According to the gene sequence of the lyase LysP53, the conventional primer design software in the prior art was adopted to design the following primer sequences:

[0054] 53P37-F: ctttaagaaggagatataccatggATGACGATGACAACAAAACGTA (as shown in SEQ ID NO.5, has an NcoI restriction site);

[0055] 53P37-R: tggtggtggtggtggtgctcgagCCCCGCCAATTCAAAGTGTGGGCT (as shown in SEQ ID NO. 6, with an XhoI restriction site).

[0056] The target gene fragment can be synthesized by a biological company, or the phage P53 genome can be used as a template to amplify the target gene. In the present invent...

Embodiment 3

[0060] Example 3 The influence of time on the activity of lyase LysP53 to crack A.baumannii WHG40137

[0061] Acinetobacter baumannii A.baumannii WHG40137 was cultivated to the logarithmic phase (OD 600nm =0.4~0.6), after the low-temperature centrifugation collects the precipitate, after washing once with Tris-HCl buffer solution, then the precipitate after washing is dissolved in the above-mentioned buffer solution to obtain the A.baumannii WHG40137 bacterial liquid, which is obtained in Example 2 The lysing enzyme LysP53 was mixed with the above-mentioned bacterial solution so that the final concentration of the lysing enzyme LysP53 was 100 μg / ml. At the same time, the mixture of the same amount of buffer and the above-mentioned bacterial solution was used as a negative control, and incubated at 37°C. Sampling at time points, and counting plate counts, the results obtained are shown in image 3 .

[0062] From image 3 It can be seen from the results that after incubation...

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Abstract

The invention discloses an antibacterial peptide P104 and lyase LysP53 with the broad-spectrum splitting activity, and application of the antibacterial peptide P104 and the lyase LysP53. The amino acid sequence of the antibacterial peptide P104 is as shown in SEQ ID NO.1, and the gene sequence of the antibacterial peptide P104 is as shown in SEQ ID NO.3; the amino acid sequence of the lyase LysP53 is as shown in SEQ ID NO.2, and the gene sequence of the lyase LysP53 is as shown in SEQ ID NO.4; the antibacterial peptide P104 and the lyase LysP53 have a relatively good splitting effect on acinetobacter baumannii, pseudomonas aeruginosa, klebsiella pneumoniae and escherichia coli in vitro, and can be used for splitting gram-negative bacteria in a broad-spectrum manner; meanwhile, the lyase LysP53 can be subjected to soluble expression in escherichia coli, and the enzyme activity is high, and thus, the antibacterial peptide P104 and the lyase LysP53 have relatively good application prospects in research and development of anti-infection drugs.

Description

technical field [0001] The invention belongs to the field of biological preparations, and in particular relates to an antimicrobial peptide P104 with broad-spectrum cleavage activity, a lyase LysP53 and applications thereof. Background technique [0002] Antibiotics were once considered to be the most powerful weapon in the treatment of bacterial infectious diseases. However, with the increasing abuse of antibiotics and the growing problem of bacterial resistance, the speed of research and development of new antibiotics is far lower than the speed of emergence of drug-resistant bacteria. The number of new antibiotics that can be used as substitutes is very small, and the cost is high, resulting in the death of a large number of patients with drug-resistant bacterial infections in clinical practice due to no cure. The preparation of phage has attracted the attention of scholars at home and abroad. [0003] Bacteriophage is a type of virus that uses microorganisms (bacteria, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/88C12N15/60C12N15/70C12N1/21A61K38/51A61P31/04C12R1/19
CPCC12N9/88C12N15/70A61P31/04A61K38/00Y02A50/30
Inventor 危宏平杨航李唱唱余军平蒋梦薇
Owner WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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