Self-assembled antibacterial peptide Lau-IL-9-C2 and application thereof

Abstract

The invention belongs to the technical field of antibacterial peptides, and particularly relates to a self-assembled antibacterial peptide Lau-IL-9-C2 and application thereof. The self-assembly antibacterial peptide Lau-IL-9-C2 provided by the invention is dendritic polypeptide with a main chain and two side chains, self-assembly driving force is provided by connecting lauric acid and depending on the strong hydrophobicity of lauric acid, and the two side chains can increase the surface polypeptide density, increase the activity and enhance the stability. Results of embodiments show that the antibacterial peptide Lau-IL-9-C2 provided by the invention can be self-assembled, is in a nano-particle state under an electron microscope, has a particle size of 10-16 nm and a critical micelle concentration of 229.1 [mu] M, is good in stability, has no obvious cytotoxicity and hemolysis effect, has relatively strong antibacterial activity on gram-negative bacteria and gram-positive bacteria, and can be used for preparing the antibacterial peptide Lau-IL-9-C2. The minimum inhibitory concentration of the compound on acinetobacter baumannii, escherichia coli, pseudomonas aeruginosa and staphylococcus aureus is 1.17-4.69 mu M.

Description

Technical Field

[0001] The invention belongs to the technical field of antimicrobial peptides, and in particular relates to a self-assembling antimicrobial peptide Lau-IL-9-C2 and an application thereof. Background Art

[0002] Microbial infections, including bacteria, fungi, viruses, etc., can cause many very serious human diseases including hepatitis, pneumonia, AIDS, tuberculosis, etc. Infectious epidemics caused by pathogenic microorganisms have always been one of the important causes of death. Since the invention of antibiotics, countless lives of patients with bacterial infections have been saved. However, with the extensive use of antibiotics, a large number of antibiotic-resistant strains have emerged.

[0003] Antimicrobial peptides (AMPs) are a class of small molecule polypeptides that can inhibit the growth of bacteria, fungi, and viruses. They are widely present in organisms and are an important part of the innate immune system. Antimicrobial peptides usually have a certain net positive charge and an amphipathic α-helical structure. On the one hand, such a structure enables antimicrobial peptides to target and bind to the negatively charged bacterial plasma membrane. On the other hand, the amphipathic characteristics can promote the interaction between antimicrobial peptides and bacterial plasma membranes, promote antimicrobial peptides to penetrate and destroy bacterial cell membranes, and ultimately lead to bacterial death. Antimicrobial peptides are not only not easy to induce bacterial resistance, but also show the advantages of broad-spectrum antibacterial, trace high efficiency, and immune regulation. However, not all small molecule polypeptides have antibacterial activity. Even if they have antibacterial activity, there are many limitations in actual production and application. The essence of antimicrobial peptides is a polypeptide, which makes them sensitive to the environment. The presence of ion concentration and enzymes will affect the structure and bactericidal effect of the peptides. In practical applications, there are instabilities, as well as unknown hemolytic activity and cytotoxicity, which seriously limit the development of antimicrobial peptides as antimicrobial drugs. There are very few antimicrobial peptide drugs that can enter phase III clinical studies. The self-assembly of peptides can improve the stability of the structure, enhance biocompatibility and increase the local concentration of action, but not all self-assembly behaviors can improve the performance of antimicrobial peptides. Summary of the invention

[0004] The purpose of the present invention is to provide a self-assembling antimicrobial peptide Lau-IL-9-C2 with good stability, no obvious cytotoxicity and hemolysis, and its application, which has strong and broad-spectrum antimicrobial activity and low minimum inhibitory concentration.

[0005] The present invention provides a self-assembling antimicrobial peptide Lau-IL-9-C2, wherein the self-assembling antimicrobial peptide Lau-IL-9-C2 comprises lauric acid, a main chain, a first side chain and a second side chain; The main chain includes the amino acid sequence shown in SEQ ID NO: 1; The first side chain and the second side chain respectively include the amino acid sequence shown in SEQ ID NO: 2; The N-terminus of the main chain is connected to the lauric acid via an amide bond; The C-terminus of the first side chain is connected to the side chain amino group of the third amino acid in the amino acid sequence shown in SEQ ID NO: 1 through an amide bond; the C-terminus of the second side chain is connected to the side chain amino group of the fourth amino acid in the amino acid sequence shown in SEQ ID NO: 1 through an amide bond.

[0006] The present invention also provides the use of the self-assembling antimicrobial peptide Lau-IL-9-C2 described in the above technical solution in the preparation of antibacterial products.

[0007] Preferably, the anti-bacterial product comprises an anti-bacterial infection product.

[0008] Preferably, the bacteria include Gram-negative bacteria and / or Gram-positive bacteria.

[0009] Preferably, the Gram-negative bacteria include Acinetobacter baumannii ( Acinetobacter baumannii )、Pseudomonas aeruginosa( Pseudomonas aeruginosa ) and Escherichia coli ( Escherichia coli )

[0010] Preferably, the Gram-positive bacteria include Staphylococcus aureus ( Staphylococcus aureus ).

[0011] Preferably, the product comprises a reagent or a drug The present invention provides an antibacterial product, which comprises the self-assembling antibacterial peptide Lau-IL-9-C2 described in the above technical solution.

[0012] Preferably, the antibacterial product comprises an antibacterial agent or an antibacterial drug.

[0013] Beneficial effects: The present invention provides a self-assembling antimicrobial peptide Lau-IL-9-C2, wherein the self-assembling antimicrobial peptide Lau-IL-9-C2 comprises lauric acid, a main chain, a first side chain and a second side chain; the main chain comprises an amino acid sequence as shown in SEQ ID NO:1; the first side chain and the second side chain respectively comprise an amino acid sequence as shown in SEQ ID NO:2; the N-terminus of the main chain is connected to the lauric acid through an amide bond; the C-terminus of the first side chain is connected to the side chain amino group of the third amino acid of the amino acid sequence as shown in SEQ ID NO:1 through an amide bond; the C-terminus of the second side chain is connected to the side chain amino group of the fourth amino acid of the amino acid sequence as shown in SEQ ID NO:1 through an amide bond. The self-assembling antimicrobial peptide Lau-IL-9-C2 of the present invention has a dendritic polypeptide with a main chain and two side chains, and provides a self-assembly driving force by connecting lauric acid and relying on its strong hydrophobicity, and the two side chains can increase the surface polypeptide density, increase activity and enhance stability. The results of the examples show that the antimicrobial peptide Lau-IL-9-C2 provided by the present invention can self-assemble and present a nanoparticle state under an electron microscope, with a particle size of 10-16 nm and a critical micelle concentration of 229.1 μM; and the self-assembled antimicrobial peptide Lau-IL-9-C2 obtained by the present invention has good stability, no obvious cytotoxicity and hemolytic effect, and has strong antibacterial activity against both Gram-negative and Gram-positive bacteria, and the minimum inhibitory concentration for Acinetobacter baumannii, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus is 1.17-4.69 μM, with a strong broad spectrum. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required to be used in the embodiments are briefly introduced below.

[0015] Figure 1 Schematic diagram of the synthesis of the self-assembling antimicrobial peptide Lau-IL-9-C2; Figure 2 The transmission electron microscopy observation results and particle size statistics of the self-assembled antimicrobial peptide Lau-IL-9-C2 are shown in Figure 1; A is a transmission electron microscopy image, and B is a particle size statistics image; Figure 3 Graph showing the results of critical micelle concentration determination of the self-assembled antimicrobial peptide Lau-IL-9-C2; wherein A is a statistical graph showing the fluorescence intensity of the gradient concentration self-assembled antimicrobial peptide Lau-IL-9-C2 at different wavelengths, and B is a graph showing the Nile red fluorescence curve of the gradient concentration self-assembled peptide Lau-IL-9-C2; Figure 4 The results of the viability test of human embryonic kidney HEK293T cells with different concentrations of self-assembled antimicrobial peptide Lau-IL-9-C2; Figure 5This is a graph showing the results of the detection of the viability of human embryonic kidney HEK293T cells by different concentrations of linear polypeptide IL-9; Figure 6 The figure shows the evaluation results of the hemolytic effect of different concentrations of the self-assembled antimicrobial peptide Lau-IL-9-C2; Figure 7 This is a graph showing the evaluation results of the hemolytic effect of different concentrations of linear polypeptide IL-9. DETAILED DESCRIPTION

[0016] The invention provides a self-assembling antimicrobial peptide Lau-IL-9-C2. The self-assembling antimicrobial peptide Lau-IL-9-C2 comprises lauric acid, a main chain, a first side chain and a second side chain; the main chain comprises an amino acid sequence as shown in SEQ ID NO:1; the first side chain and the second side chain respectively comprise an amino acid sequence as shown in SEQ ID NO:2; the N-terminus of the main chain is connected to the lauric acid through an amide bond; the C-terminus of the first side chain is connected to the side chain amino group of the third amino acid of the amino acid sequence as shown in SEQ ID NO:1 through an amide bond; and the C-terminus of the second side chain is connected to the side chain amino group of the fourth amino acid of the amino acid sequence as shown in SEQ ID NO:1 through an amide bond.

[0017] As an embodiment, the main chain is composed of a linker and a straight-chain polypeptide from the N-terminus to the C-terminus; the straight-chain polypeptide and the linker are connected by an amide bond; the amino acid sequence of the straight-chain polypeptide is shown in SEQ ID NO:3; the amino acid sequence of the linker is G. As an embodiment, the N-terminus of the connection is connected to the lauric acid by an amide bond. The present invention utilizes lauric acid, which has a suitable length, can increase activity, promote self-assembly, and will not significantly increase toxicity. Compared with other types of highly hydrophobic fatty acid chains, it is safer and has less toxicity after self-assembly of antimicrobial peptides. The self-assembling antimicrobial peptide Lau-IL-9-C2 of the present invention has a dendritic polypeptide with a main chain and two side chains, a molecular weight of 1955.41 Daltons, and all amino acids are L-type. The self-assembled antimicrobial peptide Lau-IL-9-C2 appears in a nanoparticle state under an electron microscope, with a particle size of 10-16 nm, a critical micelle concentration of 229.1 μM, good stability, no obvious cytotoxicity and hemolysis, strong antibacterial activity against both Gram-negative and Gram-positive bacteria, and a minimum inhibitory concentration of 1.17-4.69 μM against Acinetobacter baumannii, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus, with a strong broad-spectrum property.

[0018] The present invention has no strict requirements on the preparation method of the antimicrobial peptide, and conventional methods in the art can be used, such as solid phase synthesis of polypeptides. The present invention has no special requirements on the solid phase synthesis of polypeptides, and methods well known to those skilled in the art can be used.

[0019] The present invention also provides the use of the self-assembling antimicrobial peptide Lau-IL-9-C2 described in the above technical solution in the preparation of antibacterial products.

[0020] As an embodiment, the antibacterial product is an antibacterial infection product. As an embodiment, the bacteria include Gram-negative bacteria and / or Gram-positive bacteria. As an embodiment, the Gram-negative bacteria include one or more of Acinetobacter baumannii, Pseudomonas aeruginosa and Escherichia coli; as another embodiment, the Gram-positive bacteria include Staphylococcus aureus.

[0021] As an embodiment, the Acinetobacter baumannii of the present invention includes one or more of Acinetobacter baumannii ATCC 19606, 10769 and 0357; as another embodiment, the Acinetobacter baumannii of the present invention is Acinetobacter baumannii ATCC 19606. The present invention is described in the embodiments using Acinetobacter baumannii ATCC 19606, 10769 and 0357 as examples, but they should not be understood as the entire protection scope of the present invention.

[0022] As an embodiment, the Escherichia coli of the present invention includes one or more of Escherichia coli ATCC 8739, 0894 and 5017; as another embodiment, the Escherichia coli of the present invention includes Escherichia coli ATCC 8739. The present invention is described by taking Escherichia coli ATCC 8739, 0894 and 5017 as examples in the embodiments, but they cannot be understood as the entire protection scope of the present invention.

[0023] As an embodiment, the Pseudomonas aeruginosa described in the present invention includes one or more of Pseudomonas aeruginosa ATCC 27853, 90068 and 17068; as another embodiment, the Pseudomonas aeruginosa described in the present invention is Pseudomonas aeruginosa ATCC 27853. The present invention is described in the embodiments using Pseudomonas aeruginosa ATCC 27853, 90068 and 17068 as examples, but they should not be construed as the entire protection scope of the present invention.

[0024] As an embodiment, the Staphylococcus aureus of the present invention includes one or more of Staphylococcus aureus ATCC 6538, 220823 and 15775; as another embodiment, the Staphylococcus aureus of the present invention is Staphylococcus aureus ATCC 6538. The present invention is described in the embodiments using Staphylococcus aureus ATCC 6538, 220823 and 15775 as examples, but they cannot be understood as the entire protection scope of the present invention.

[0025] In the present invention, the strains corresponding to numbers 10769, 0357, 0894, 5017, 90068, 170682, 20823 and 15775 are respectively known strains in the art, deposited in the Institute of Medical Biology, Chinese Academy of Medical Sciences, and disclosed in the prior art Development of α-Helical Antimicrobial Peptides with Imperfect Amphipathicityfor Superior Activity and Selectivity. J Med Chem. 2024 Nov 14;67(21):19561-19572.

[0026] As an embodiment, the product includes a reagent or a drug.

[0027] The present invention provides an antibacterial product, which includes the self-assembling antibacterial peptide Lau-IL-9-C2 described in the above technical solution; the antibacterial product includes an antibacterial agent or an antibacterial drug. As an embodiment, the minimum inhibitory concentration of the antibacterial peptide in the antibacterial product is ≥1.17 μM; as another embodiment, the minimum inhibitory concentration of the antibacterial peptide in the antibacterial product is 1.17~4.69 μM; as another embodiment, the minimum inhibitory concentration of the antibacterial peptide in the antibacterial product is 1.17~2.34 μM; as another embodiment, the minimum inhibitory concentration of the antibacterial peptide in the antibacterial product is 1.17 μM.

[0028] To further illustrate the present invention, a self-assembling antimicrobial peptide Lau-IL-9-C2 and its application provided by the present invention are described in detail below in conjunction with the accompanying drawings and examples, but they should not be construed as limiting the scope of protection of the present invention.

[0029] Example 1 Commissioned Jier Biochemical (Shanghai) Co., Ltd. to synthesize peptides through solid phase synthesis. Figure 1A dendritic polypeptide is synthesized according to a flow chart, and is desalted and purified by HPLC reverse phase column chromatography to obtain the dendritic polypeptide, which is named Lau-IL-9-C2; the dendritic polypeptide Lau-IL-9-C2 has a main chain and two side chains, and the N-terminus of the main chain is connected to lauric acid (Lau) through an amide bond; wherein the main chain amino acid sequence is GIKKGIKKLL (SEQ ID NO: 1), which consists of 10 amino acids; the amino acid sequences of the two side chains are both KGIKKLL (SEQ ID NO: 2), which consist of 7 amino acids, and are respectively connected to the side chain amino groups of the third lysine (K) and the fourth lysine (K) of the main chain through amide bonds; the third lysine is the third amino acid residue from the N-terminus to the C-terminus of the main chain, and the fourth lysine is the fourth amino acid residue from the N-terminus to the C-terminus of the main chain; the molecular weight of Lau-IL-9-C2 is 2841.9 Daltons, and all amino acids are L-type.

[0030] Comparative Example 1 We commissioned Gill Biochemical (Shanghai) Co., Ltd. to synthesize a linear polypeptide IL-9 with only 9 amino acids through peptide solid phase synthesis. Its amino acid sequence is IKKGIKKLL (SEQ ID NO: 3), all of which are L-type amino acids.

[0031] Test Example 1 Transmission electron microscopy observation (1) Drop the solution containing the dendritic peptide Lau-IL-9-C2 obtained in Example 1 onto glossy paper, clamp a 300-mesh copper mesh, with the carbon film facing down, cover and adsorb for 5 minutes, and absorb the excess liquid; after uranium staining (2%wt uranyl acetate) for 2 minutes, absorb the excess dye, dry it, and observe and take pictures using a transmission electron microscope (model JEM-1400 Plus, JEOL Ltd.). The results showed that the dendritic peptide Lau-IL-9-C2 was in the form of nanoparticles under the transmission electron microscope, indicating that the dendritic peptide Lau-IL-9-C2 can form nanoparticles by self-assembly, which is a self-assembling peptide Lau-IL-9-C2 ( Figure 2 Middle A).

[0032] (2) Data processing: Image J software was used to calculate the particle size of nanoparticles, and GraphPad Prism was used to plot the particle size distribution. The results showed that the particle size of the self-assembling peptide Lau-IL-9-C2 was 10-16 nm ( Figure 2 Middle B).

[0033] Test Example 2 Critical micelle concentration (CMC) determination (1) Add 100 μL of gradient concentrations of self-assembling peptide Lau-IL-9-C2 solution to a 96-well fluorescent ELISA plate with a black bottom, and then add 100 μL of 25 μM Nile red solution to each well. After mixing, place at room temperature for 30 minutes, and scan the emission spectrum of the solution with a multifunctional ELISA reader ( Figure 3 Middle A).

[0034] The buffer solution used to prepare the Nile red solution and the gradient concentration self-assembling polypeptide Lau-IL-9-C2 solution was 10 mM phosphate buffer solution (pH 7.4); the fluorescence spectrum scanning conditions were: excitation wavelength of 550 nm, emission wavelength of 600-700 nm, and bandwidth of 1 nm.

[0035] (2) Data processing: The logarithmic value of the concentration of the self-assembling peptide Lau-IL-9-C2 was used as the horizontal axis, and the maximum fluorescence intensity of Nile red at each concentration was used as the vertical axis to plot the fluorescence curve of Nile red in the presence of gradient concentrations of the self-assembling peptide Lau-IL-9-C2 (excitation wavelength of 550 nm, emission wavelength of 630 nm). The peptide concentration corresponding to the inflection point of the fluorescence curve is the minimum concentration at which the self-assembling peptide Lau-IL-9-C2 begins to form micelles in aqueous solution, that is, the CMC value ( Figure 3 Middle B).

[0036] according to Figure 3 It can be seen that the CMC value of the self-assembling peptide Lau-IL-9-C2 is 229.1 μM. Nile red is a lipophilic fluorescent dye that produces strong fluorescence after being inserted into the hydrophobic pocket of the micelle. When the concentration of the self-assembling peptide Lau-IL-9-C2 is higher than the CMC, the fluorescence of Nile red in the solution can be significantly increased, indicating that the self-assembling peptide Lau-IL-9-C2 self-assembles in the aqueous solution to form amphiphilic micelles with hydrophilic periphery and hydrophobic interior.

[0037] Test Example 3 Antimicrobial activity analysis 1. Test strains (1) Acinetobacter baumannii: ATCC 19606, 10769 and 0357; (2) Escherichia coli: ATCC 8739, 0894 and 5017; (3) Pseudomonas aeruginosa: ATCC 27853, 90068 and 17068; (4) Staphylococcus aureus: ATCC 6538, 220823 and 15775.

[0038] 2. Minimum inhibitory concentration (MIC) test First, the test strain was inoculated onto the LB solid plate. After the colonies grew, a single colony was picked and transferred to the LB liquid medium. The culture was placed at 37°C and 180 rpm for 5 hours. The OD of the bacterial solution was measured under a UV spectrophotometer. 600 , according to 1OD 600 =1×10 9 CFU / mL ratio, dilute the bacterial solution to 2×10 5 CFU / mL concentration; 100 μL of diluted bacterial solution was added to each sterile 96-well plate, and then 100 μL of the sample to be tested diluted with physiological saline in a gradient manner was added to each well, and the mixture was evenly mixed by pipetting, and then placed in a 37°C constant temperature incubator for slow shaking and culture overnight; the sample to be tested was Lau-IL-9-C2 obtained in Example 1 and the linear polypeptide IL-9 obtained in Comparative Example 1; the concentration of the diluted sample to be tested was 0-200 μg / mL; after constant temperature culture, the bacterial solution was measured at OD using an ELISA instrument 600 The absorbance value at nm was taken as the MIC value according to the average of the sample concentrations of the wells where no bacterial growth was detected and the adjacent wells. The results are shown in Table 1.

[0039] Table 1 Minimum inhibitory concentration values ​​of the tested samples for the tested strains

[0040] It can be seen from Table 1 that Lau-IL-9-C2 obtained in Example 1 exhibited significant antibacterial effects on all tested strains, with MIC values ​​ranging from 1.17 to 4.69 μM. Compared with the linear polypeptide IL-9 in Comparative Example 1, the antibacterial activity changed from none (MIC>50 μM) to some.

[0041] Test Example 4 Plasma stability analysis 1. Test strains Escherichia coli: ATCC 8739; Staphylococcus aureus: ATCC 6538; Acinetobacter baumannii: ATCC 19606; Pseudomonas aeruginosa: ATCC 27853.

[0042] 2. Take a 1.5mL EP tube and add 100μL 15mg / mL EDTA-K2 anticoagulant. Remove the mouse eyeball and collect blood into the above tube. Immediately invert to mix. Centrifuge at 4℃ and 3500rpm for 10min. Carefully aspirate the supernatant to obtain mouse plasma.

[0043] 3. The Lau-IL-9-C2 obtained in Example 1 was used as the test sample, dissolved in sterile physiological saline, and the final concentration was 400 μg / mL; the test sample solution was mixed with mouse plasma in a volume ratio of 1:1, and incubated in a 37°C incubator. 200 μL of the sample was taken out at 0, 0.5, 1, 2, 4, 6, and 8 hours of incubation for MIC experimental detection to see the change in antibacterial activity after incubation with plasma. The MIC test method is as described in Example 2, and the specific test results are shown in Table 2.

[0044] Table 2 Effect of plasma on the antibacterial activity of Lau-IL-9-C2 (MIC, μM)

[0045] According to Table 2, the antibacterial activity of Lau-IL-9-C2 against the four standard strains did not change after incubation with plasma for 8 h, indicating that it has excellent plasma stability.

[0046] Test Example 5 Cytotoxicity assay Human embryonic kidney cells (HEK293T) were routinely cultured in DMEM medium (containing 10% fetal bovine serum and 1% double antibody). When the cells grew to 80% of the culture flask, they were washed three times with phosphate buffer, then digested with 0.25% trypsin, and diluted to 5×10 5 100 μM / mL cell suspension was added to each well of a sterile 96-well plate, and the culture was continued overnight. On the second day, different concentration gradients of the samples to be tested (final concentration of 0-100 μM) or the same volume of DMEM culture medium were added, and 3 replicates were set for each sample concentration. After culturing for 24 hours, 15 μL of 5 mg / mL MTT solution was added to each well, and the culture was continued for 4 hours in the dark. Then, the culture medium in the plate wells was discarded, and 200 μL DMSO was added to each well. The culture plate was placed on a shaker and gently shaken for 10 minutes to dissolve the crystals. Then, the absorbance value of each well was measured at 450 nm using an enzyme reader. The results are shown in Figure 2. Figure 4 and Figure 5 As shown; wherein, the sample to be tested is Lau-IL-9-C2 obtained in Example 1 and the linear polypeptide IL-9 obtained in Comparative Example 1.

[0047] according to Figure 4 and Figure 5 It can be seen that the linear polypeptide IL-9 obtained in Comparative Example 1 has no toxicity to human keratinocytes HaCaT within the tested concentration range; the Lau-IL-9-C2 obtained in Example 1 has no toxicity to human keratinocytes HaCaT within the concentration range of 50 μM, and only exhibits significant cytotoxicity at a concentration of 100 μM (** P<0.01). The present invention realizes the antibacterial activity of the linear polypeptide IL-9 from no (MIC>50μM) to yes through self-assembly, while having no cytotoxicity and good safety.

[0048] Test Example 6 Hemolysis assay 1. Mix fresh human whole blood and Aldrich solution in a volume ratio of 1:1, centrifuge at 1000rpm for 5min, discard the supernatant, wash the obtained red blood cells with physiological saline, repeat 3 times until the supernatant no longer appears red, and obtain the washed red blood cells.

[0049] 2. Dilute the washed red blood cells to 1×10 7 The red blood cell suspension was incubated with different concentrations (0-100 μM) of the test sample dissolved in physiological saline at 37°C for 30 min, and then centrifuged at 1000 rpm for 5 min. The absorbance of the supernatant was detected at 540 nm. The positive control used the same volume of Triton X-100 solution (PC), and the volume concentration of the Triton X-100 solution was 10%; the negative control used physiological saline; wherein the test sample was Lau-IL-9-C2 obtained in Example 1 and the linear polypeptide IL-9 obtained in Comparative Example 1.

[0050] 3. Data processing: Detect the absorbance at 540nm, define the hemolysis rate of the positive control (PC group) as 100%, and calculate the hemolysis rate of the sample group to be tested relative to the PC group according to the following formula. The results are as follows: Figure 6 and Figure 7 shown.

[0051] Hemolysis rate (%) = (positive control OD 540 Value - negative control OD 540 value) / (OD of the sample group to be tested 540 Value - negative control OD 540 value) × 100%.

[0052] according to Figure 6 and Figure 7 It can be seen that the linear polypeptide IL-9 obtained in Comparative Example 1 and Lau-IL-9-C2 obtained in Example 1 show higher safety to human red blood cells and have no significant hemolytic effect within a concentration range of 100 μM. The present invention achieves the antibacterial activity of the linear polypeptide IL-9 from no (MIC>50 μM) to yes by self-assembly, without significant hemolytic effect and good safety.

[0053] Based on the above content, it can be seen that the self-assembling antimicrobial peptide Lau-IL-9-C2 provided by the present invention has strong antibacterial activity against both Gram-negative and Gram-positive bacteria, good stability, strong broad spectrum, extremely high safety, and no cytotoxicity or hemolytic effect.

[0054] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A self-assembling antimicrobial peptide Lau-IL-9-C2, characterized in that: The self-assembling antimicrobial peptide Lau-IL-9-C2 comprises lauric acid, a main chain, a first side chain and a second side chain; The main chain includes the amino acid sequence shown in SEQ ID NO: 1; The first side chain and the second side chain respectively include the amino acid sequence shown in SEQ ID NO: 2; The N-terminus of the main chain is connected to the lauric acid via an amide bond; The C-terminus of the first side chain is connected to the side chain amino group of the third amino acid in the amino acid sequence shown in SEQ ID NO: 1 through an amide bond; the C-terminus of the second side chain is connected to the side chain amino group of the fourth amino acid in the amino acid sequence shown in SEQ ID NO: 1 through an amide bond.

2. Use of the self-assembling antimicrobial peptide Lau-IL-9-C2 according to claim 1 in the preparation of antibacterial products.

3. The use according to claim 2, characterized in that: The anti-bacterial products include anti-bacterial infection products.

4. The use according to claim 2, characterized in that: The bacteria include Gram-negative bacteria and / or Gram-positive bacteria.

5. The use according to claim 4, characterized in that: The Gram-negative bacteria include Acinetobacter baumannii ( Acinetobacter baumannii )、Pseudomonas aeruginosa( Pseudomonas aeruginosa ) and Escherichia coli ( Escherichia coli ) 6. The use according to claim 4, characterized in that: The Gram-positive bacteria include Staphylococcus aureus ( Staphylococcus aureus ).

7. The use according to any one of claims 2 to 6, characterized in that: The product includes a reagent or a drug.

8. An antibacterial product, characterized in that: The antibacterial product comprises the self-assembling antimicrobial peptide Lau-IL-9-C2 according to claim 1.

9. The antibacterial product according to claim 8, characterized in that The antibacterial product includes an antibacterial agent or an antibacterial drug.

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