CN113248572A

CN113248572A - Anti-multidrug-resistant bacteria cyclopeptide and application thereof

Info

Publication number: CN113248572A
Application number: CN202110477846.4A
Authority: CN
Inventors: 王远强; 何清秀; 李广平; 赵丽楠; 庹燕; 刘周; 李跃鹏
Original assignee: Chongqing University of Technology
Current assignee: Chongqing University of Technology
Priority date: 2021-04-30
Filing date: 2021-04-30
Publication date: 2021-08-13
Anticipated expiration: 2041-04-30
Also published as: CN113248572B

Abstract

The invention discloses an anti-multiple drug-resistant bacterial cyclopeptide and application thereof, which is characterized in that the amino acid sequence is any one of the following six types: R-R-W-W-W-R, R-W-R-W-R-W, R-R-W-W-W-R-R, R-R-W-W-R-R-W, R-R-W-W-W-W-R-R, R-R-W-W-R-R-W. The antibacterial cyclopeptide is obtained by cyclization of any one of the amino acid sequences. The invention provides 6 novel antibacterial peptides of artificially designed cations. The antibacterial peptides can be synthesized by Fmoc solid phase chemistry. The cationic antibacterial peptide has broad-spectrum killing activity on multiple drug-resistant acinetobacter baumannii, staphylococcus aureus and escherichia coli, has no toxic action on animal and plant cells, and has strong operability and low cost.

Description

Anti-multidrug-resistant bacteria cyclopeptide and application thereof

Technical Field

The invention relates to a cyclic peptide for resisting multiple drug-resistant bacteria and application thereof, and relates to the field of cyclic peptide.

Background

With the wide clinical application of antibiotics, multidrug-resistant bacteria are also continuously appearing and increasing. In recent years, the emergence of multi-drug resistant gram-negative bacteria (escherichia coli, pseudomonas aeruginosa, acinetobacter baumannii) and gram-positive bacteria (staphylococcus aureus), which are the most common cause of nosocomial infections, has been an increasing problem and challenge worldwide. In the face of increasingly serious drug resistance problems, however, enough drugs for solving the problems have not been developed yet, polymyxins (such as colistin) become the last effective means for treating drug-resistant bacterial infections again, and therefore, the development of novel antibacterial drugs is urgently needed.

The antibacterial peptide (AMP) is a short peptide existing in all organisms, has the characteristics of cationic property and amphipathy, has various structures, is an important component of a natural immune system of the organisms to resist the invasion of pathogenic bacteria, and keeps stable and efficient in the biological evolution process. The antibacterial peptide has good water solubility and high thermal stability, has biological effects of broad-spectrum antibiosis, antifungal, antivirus, antitumor cell and the like, and can also be used as an immunomodulator to play a plurality of immune effects in organisms, such as acting as a chemotactic factor, inducing the generation of the chemotactic factor, promoting wound healing, regulating dendritic cell and cellular immune response and the like. Most of antibacterial peptides kill bacteria through a unique cell membrane damage mechanism, the action mode is different from that of the currently clinically used antibiotics, pathogenic bacteria hardly generate drug resistance of the antibacterial peptides through changing cell membrane components, and therefore the antibacterial peptides show good application prospects in the aspect of treating infection caused by drug-resistant bacteria, are widely concerned in recent years, and become a hotspot for research and development of novel antibacterial drugs.

In order to find antimicrobial peptides from polypeptides, one initially studied polypeptides by conducting experiments and identified them by observing whether they have antimicrobial activity. In this way, although it can be accurately determined whether the polypeptide has an antimicrobial effect. However, the experimental procedure is cumbersome, takes a long time, requires a large amount of money, and cannot predict the activity of the antimicrobial peptide. With the development of high-throughput proteomics, the number of protein and polypeptide sequences has increased rapidly. It is difficult to identify effective antimicrobial peptides from a large number of peptide samples and to experimentally predict their antimicrobial activity. Therefore, there is a need to find additional methods to identify and predict effective antimicrobial peptides. With the continuous development of bioinformatics, computational methods are used for screening and activity prediction, which effectively solve various disadvantages of experimental methods.

Disclosure of Invention

Aiming at the technical problems, the invention aims to provide an antibacterial cyclic peptide with multiple drug resistance and also provides application of the antibacterial cyclic peptide, which has strong antibacterial activity, low cost and low hemolytic toxicity.

In order to achieve the purpose, the technical scheme of the invention is as follows: the cyclic peptide for resisting multiple drug resistance bacteria is characterized in that the amino acid sequence is any one of the following six types:

CPeptide-A：R-R-W-W-W-R(Arg-Arg-Trp-Trp-Trp-Arg)

CPeptide-B:R-W-R-W-R-W(Arg-Trp-Arg-Trp-Arg-Trp)

CPeptide-C:R-R-W-W-W-R-R(Arg-Arg-Trp-Trp-Trp-Arg-Trp)

CPeptide-D:R-R-W-W-R-R-W(Arg-Arg-Trp-Trp-Arg-Arg-Trp)

CPeptide-E:R-R-W-W-W-W-R-R(Arg-Arg-Trp-Trp-Trp-Trp-Arg-Arg)

CPeptide-F:R-R-W-W-R-R-R-W(Arg-Arg-Trp-Trp-Arg-Arg-Arg-Trp)。

the structural formulas after cyclization are respectively as follows:

CPeptide-A:

CPeptide-B:

CPeptide-C:

CPeptide-D:

CPeptide-E:

CPeptide-F:

the invention designs cationic cyclic peptide based on sequence and structure analysis of natural antibacterial peptide. Bovine lactoferrin LfcinB6_4-9(RRWQWR) is used as a template, combines the characteristics of polypeptide sequences, virtually combines, designs and screens to obtain an optimal amino acid combination sequence, and can better form a restrictive conformation based on cyclic peptide, thereby enhancing the binding capacity with a target molecule and the selectivity to a corresponding receptor, and having the characteristics of easy penetration of bacterial membranes, low toxicity, endopeptidase and exopeptidase hydrolysis resistance and the like. On the basis, amino acid sequence design and transformation are carried out and cyclized to construct an antibacterial cyclic peptide library. And (3) taking the BamA protein as a target, and virtually screening by molecular docking to obtain the potential antibacterial cyclic peptide. The 6 straight-chain peptides which are designed and screened are respectively 'RRWWWR', 'RWRWRWRWRRW', 'RRWWWRR', 'RRWWRRW', 'RRWWWWWRR' and 'RRWWRRRW', and the experimental research shows that the peptides have stronger antibacterial ability and small hemolytic toxicity after cyclization.

The preparation method of the novel cationic antibacterial cyclic peptide is a solid-phase synthesis method of polypeptide, firstly resin is required to be swelled, and then C-terminal carboxyl of first amino acid and activity on the resin are combinedAnd (3) carrying out site chlorine reaction, carrying out dehydration condensation to connect second amino acid after the first amino acid is connected on the resin, and removing Fmoc protection after condensation is finished. Repeating the operation according to the designed amino acid sequence, sequentially completing the rest amino acids, and finally cutting the fully-protected linear polypeptide from the resin by using a cutting reagent. Reacting the fully-protected linear peptide with a condensing agent DIC/Cl-HOBt, adding water to precipitate a solid after the reaction is finished to obtain the protected cyclic peptide, and adding 95% TFA/H₂And cracking O at room temperature for 2 hours to obtain a cyclic peptide primary product.

In order to deeply research the relationship between the structure and the function of the bioactive antibacterial peptide, the sterilization and bacteriostasis capacities of the polypeptide are detected by an agar perforation method and a minimum bacteriostasis concentration experiment, and the clinical common antibiotic imipenem is used as a positive control, and the result shows that the sterilization activity and the bacteriostasis capacity of the antibacterial cyclic peptide are equivalent to those of imipenem.

The detection of the cytotoxicity of the antibacterial cyclic peptide in normal cells is a necessary measure which can be clinically used, and the antibacterial effect of the cationic antibacterial cyclic peptide is that the cationic antibacterial cyclic peptide has electropositivity and can generate electrostatic attraction with negatively charged phosphatidyl glycerol and cardiolipin in a cell membrane, so that the structure of the cell membrane of bacteria is damaged and the bacteria is prevented from being bacteria. However, since phosphatidylserine and phosphatidylinositol exist in the cell membrane of eukaryotic cells, and both of them have negative charges, there is a possibility that the antibacterial cyclic peptide is bound to the cell membrane of eukaryotic cells, which may cause cytotoxicity. Therefore, the hemolytic toxicity of the polypeptide needs to be detected, and experiments show that the hemolytic rate of the antibacterial peptide is still very low at high concentration, which proves that the hemolytic toxicity of the antibacterial cyclic peptide is very low.

The second object of the present invention is achieved by: an application of the multi-drug resistant antibacterial cyclopeptide in antibiosis.

An application of the multi-drug resistant bacteria resisting cyclic peptide in resisting multi-drug resistant acinetobacter baumannii, escherichia coli and staphylococcus aureus.

Has the advantages that: the invention provides 6 novel antibacterial cyclic peptides of artificially designed cations. The antibacterial cyclic peptide can be synthesized by adopting Fmoc solid phase chemical method. The cationic antibacterial cyclic peptide has broad-spectrum killing activity on multiple drug-resistant acinetobacter baumannii, staphylococcus aureus and escherichia coli, has no toxic action on animal and plant cells, and has strong operability and low cost.

Drawings

FIG. 1 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-A.

FIG. 2 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-B.

FIG. 3 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-C.

FIG. 4 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-D.

FIG. 5 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-E.

FIG. 6 is a mass spectrum of the antimicrobial cyclic peptide CPeptide-F.

Detailed Description

The invention is further illustrated by the following examples in conjunction with the accompanying drawings:

the target products CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-E and CPeptide-F are artificially synthesized into the antibacterial peptide according to a standard Fmoc solid phase program:

CPeptide-A:(Arg-Arg-Trp-Trp-Trp-Arg),

CPeptide-B:(Arg-Trp-Arg-Trp-Arg-Trg),

CPeptide-C:(Arg-Arg-Trp-Trp-Trp-Arg-Arg),

CPeptide-D:(Arg-Arg-Trp-Trp-Arg-Arg-Trp),

CPeptide-E:(Arg-Arg-Trp-Trp-Trp-Trp-Arg-Arg),

CPeptide-F:(Arg-Arg-Trp-Trp-Arg-Arg-Arg-Trp),

the synthesized product was purified by reverse phase liquid chromatography (Vydac 218TP1022 column 2.2 × 25cm), eluted with acetonitrile/water system, and then analyzed by mass spectrometry. The sequence of the prepared CPeptide-A polypeptide is as follows: R-R-W-W-W-R (Arg-Arg-Trp-Trp-Trp-Arg), the CPeptide-B polypeptide sequence is R-W-R-W-R-W (Arg-Trp-Arg-Trp-Arg-Trg), the CPeptide-C polypeptide sequence is R-R-W-W-R-R (Arg-Arg-Trp-Trp-Trp-Arg-Arg), the CPeptide-D polypeptide sequence is R-R-W-W-R-W (Arg-Arg-Trp-Trp-Arg-Arg-Trp), and the CPeptide-E polypeptide sequence is R-R-W-W-R-R (Arg-Arg-Trp-Trp-Arg-Arg-Trp), and the CPeptide-E polypeptide sequence is R-R-W-W-W-W-R (Arg-Arg-Trp-Trp-Trp- Trp-Arg-Arg), and the CPeptide-F polypeptide sequence is R-R-W-W-R-R-R-W (Arg-Arg-Trp-Trp-Arg-Arg-Trp).

Example 1

1. CPeptide-A (Arg-Arg-Trp-Trp-Trp-Arg), cyclic peptide synthesis.

Fmoc solid-phase polypeptide synthesis is adopted, 2-CTC resin is used as a carrier, full-protection peptide linear peptide is synthesized firstly, cyclization is carried out in a liquid phase, and finally, deprotection is carried out by TFA liquid, and ether precipitation is carried out to obtain crude peptide.

1g of the initial resin was swollen with 10ml of DCM at room temperature for 30 min;

first amino acid coupling procedure:

weighing protected amino acid Fmoc-Arg (Pbf) -OH with the total resin substitution value of 3eq, adding the protected amino acid Fmoc-Arg (Pbf) -OH into a DCM solution, adding DIEA with the total resin substitution value of 9eq for dissolution, adding the dissolved clear solution into resin for coupling reaction for 3 hours, discharging waste liquid, and washing with DMF for 3 times.

Second amino acid coupling procedure:

removing Fmoc protecting groups: adding 5ml of 20% PPD/DMF reagent into the reaction tube for reaction for 5min, discharging the reaction solution, and adding 5ml of 20% PPD/DMF solution again for reaction for 25 min;

washing after deprotection: washing with DMF solution for 8 times, each time for 3min, and each time using 5 ml; after washing, taking a small amount of resin, and carrying out color development detection on the resin by using a bromophenol blue solution, wherein the resin is in a deep color;

coupling Fmoc-AA-OH: weighing 3eq of amino acid to be coupled and protected and 3eq of Cl-HOBt, adding 4ml of DMF for dissolving, adding 3eq of DIC after dissolving, oscillating and mixing for 1-2min, adding the solution into a reaction tube, and reacting with the deprotected resin at room temperature for 1.5 h;

washing after coupling: the resin was washed 5 times with DMF in 3min each time, 5ml each time. After washing, taking a small amount of resin, and carrying out color development detection by using bromophenol blue solution, wherein the resin is almost colorless.

Coupling all the protected amino acids in turn according to a second amino acid coupling mode, removing the Fmoc protecting group of the last protected amino acid, washing with DMF for 4 times, then washing with DCM for 5 times, and vacuum-drying under reduced pressure.

The resin was cleaved 3-5 times with 5 volumes by weight of TFA/DCM (concentration about 2%) solution for each timeFor 3 minutes. The lysate was immediately taken as 10% NaHCO₃The solution was adjusted to neutral. And combining the lysates for multiple times, concentrating under vacuum and reduced pressure, separating out a solid, filtering and washing with water. And drying the solid under vacuum and reduced pressure to obtain the fully-protected linear peptide.

The fully protected linear peptide was dissolved in THF or DCM, and if the solubility was poor, a little DMF was added to aid the dissolution. The final concentration of the fully protected linear peptide is controlled to be below 5 mg/ml. Adding a condensing agent DIC/Cl-HOBt for reaction, and monitoring the reaction process by a liquid phase.

After the reaction, the solvent was distilled off, and water was added to precipitate a solid. And washing the solid with water and drying to obtain the fully-protected cyclic peptide.

The fully protected cyclic peptide was treated with 95% TFA/H₂And (3) cracking the O at room temperature for 2h, adding methyl tert-butyl ether to separate out a crude product, washing, drying and inspecting.

The crude peptide was purified using a preparative liquid phase.

Firstly, dissolving crude peptide with 30-50mL of 50% acetonitrile solution, carrying out ultrasonic treatment for 2min, filtering the dissolved solution with a filter membrane, taking 3uL of solution, and analyzing the crude peptide by using analytical grade HPLC. Gradient elution is carried out by using water and acetonitrile as mobile phases for 30min, HPLC is firstly balanced for 5min, and then sample injection is carried out. And dissolving the sample, injecting the sample, collecting the sample, and preparing the dissolved sample for injection. Preparative HPLC equilibrated for 10min with an initial gradient: water 95%, acetonitrile 5%, end gradient: 25% of water, 75% of acetonitrile and 40min of gradient time. The sample from the detector is collected.

The preparation of other antibacterial cyclopeptides is the same as that of CPeptide-A (Arg-Arg-Trp-Trp-Trp-Arg), and the structural formulas of the finally prepared antibacterial cyclopeptides are respectively as follows:

CPeptide-A:

CPeptide-B:

CPeptide-C:

CPeptide-D:

CPeptide-E:

CPeptide-F:

identification of antibacterial cyclic peptides

After mass spectrum analysis, the molecular weights displayed in the mass spectrum of the prepared antibacterial cyclic peptides CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-E and CPeptide-F are 1027.20, 1027.35, 1183.20, 1183.40, 1369.80 and 1339.60 respectively, and the theoretical values calculated by polypeptide sequences are 1027.20, 1027.20, 1183.39, 1183.39, 1369.60 and 1339.58. The prepared polypeptide is proved to be the designed CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-E, CPeptide-F antibacterial cyclic peptide. And (4) identifying qualified antibacterial cyclic peptide products for later use. The control antimicrobial peptide LfcinB64-9 was prepared using a synthetic method similar to that used for the CPeptide-A antimicrobial peptide.

Experimental example 1 measurement of bactericidal Activity of cationic antibacterial Cyclic peptide

The various strains used in the following examples were purchased from the china institute for biologicals assay.

The bactericidal activity of the cationic antibacterial cyclic peptide is detected by an agar perforation method, and the bactericidal activity of CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-E, CPeptide-F in the invention is evaluated by taking the cationic antibacterial peptide LfcinB64-9 RRWQWR synthesized by a solid phase chemical method as a control.

The bactericidal activity of the antimicrobial peptide is determined according to the following steps:

recovering strains: inoculating drug-resistant acinetobacter baumannii into NA nutrient agar culture medium, streaking, and culturing in a constant-temperature incubator at 37 ℃ for 16-20 hours.

And (3) strain culture: selecting single colony, placing in 100ml MHB culture medium for culturing at the same optimum growth temperature of 37 ℃ and the shaking table rotation speed of 160 r/min, and performing shaking culture (16-20h) to enable the growth state of bacteria to reach logarithmic phase.

Preparing a bacterial suspension: the concentration of bacteria is generally measured by a turbidimetric tube (McLeod) with a turbidity of about 0.5 McLeod, at which the bacterial colony count is about 1.5X 10⁸cfu/ml, then diluted to 10 at a ratio of 1:1000⁵-10⁶cfu/ml bacterial suspension.

And (3) antibacterial experiment: uniformly coating the diluted bacterial suspensions on 25ml of NA culture medium according to the amount of 0.1ml per plate; and (5) after the bacterial liquid is solidified, punching (the diameter is 9 mm). Adding 50ul (1mg/ml, 0.5mg/ml, 0.25mg/ml) of imipenem to the positive control; negative control, adding 50ul deionized water; 50ul (1mg/ml, 0.5mg/ml, 0.25mg/ml) of the antimicrobial peptide solution was added to each of the other wells. And (3) performing bacterial culture in a constant-temperature incubator at 37 ℃, and measuring the size of a bacteriostatic zone of the bacteria after 16h, so that the size of the bacteriostatic activity of the bacteria can be preliminarily determined, and three groups of parallel experiments are performed.

TABLE 1 inhibition zone diameter (mm) of cyclic peptides of different concentrations against multiple drug-resistant strains of Acinetobacter baumannii

Note: the diameter of the zone of inhibition is less than or equal to 9mm, and the product is judged to have no inhibition effect

The results show that the bactericidal capacity of the cationic antibacterial peptide is obviously superior to that of the control antibacterial peptide LfcinB6_4-9(RRWQWR), especially the antibacterial effect of the antibacterial cyclopeptide CPeptide-D is obviously higher than that of other antibacterial cyclopeptides, and is close to that of imine cultureAnd (5) south.

Experimental example 2 measurement of bacteriostatic Activity of cationic antibacterial Cyclic peptide

The minimum bacteriostatic ability of the cationic antibacterial cyclic peptide is determined, and the cationic antibacterial peptide LfcinB6 synthesized by the solid phase chemical method is used_4-9RRWQWR was used as a control to evaluate the bacteriostatic ability of CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-E, CPeptide-F of the present invention.

The antibacterial activity of the antibacterial cyclic peptide is determined according to the following steps:

the log phase grown bacteria were collected, centrifuged at 8000 rpm for 2min at 4 ℃, washed 3 times with physiological saline, and fresh broth was added to give a bacterial suspension concentration of 2.0X 105 cfu/mL. 50uL of the bacterial suspension (100 uL PBS in the peripheral wells) was added to the experimental wells of the 96-well cell culture plate, and 50uL of peptide solutions (imipenem solution) with different concentrations were added, so that the final concentration (ug/mL) of the peptide solution (imipenem solution) in each well of the horizontal row was: 512. 256, 128, 64, 32, 16, 12, 4. The PBS buffer solution with the same volume is used as a growth control group, three parallel groups are arranged in each group, after a cell culture plate is covered, the cell culture plate is placed in a biochemical incubator at 37 ℃ for culture for 12h, and the bacterial growth condition (OD600 nm) in each hole is determined through full-automatic enzyme mapping. The minimum Inhibitory concentration mic (minimum Inhibitory concentrations) is defined as the peptide concentration in the well where bacterial growth is completely inhibited.

TABLE 26 comparison of the Minimum Inhibitory Concentration (MIC) of the antibacterial Activity of the antimicrobial Cyclic peptides against different bacteria

The smaller the minimum inhibitory concentration value in the table, the stronger the antibacterial ability of the antibacterial cyclic peptide. As can be seen from the above table, the six antibacterial cyclic peptides of the present invention have lower minimum inhibitory concentrations and MIC ratios of LfcinB6 as compared to the control peptide_4-9Are much smaller, illustrating the antimicrobial capacity of the cationic antimicrobial cyclic peptides of the present inventionFar stronger than the control antibacterial peptide.

EXAMPLE 3 in vitro assay for hemolytic Activity

This example was used to determine the hemolytic rate of cationic antimicrobial cyclic peptide to sheep red blood cells, and used cationic antimicrobial peptide LfcinB64-9 RRWQWR synthesized by solid phase chemistry as a control. The blood samples used were obtained from defibrinated sheep blood.

The detection steps of the hemolysis rate of the cationic antibacterial cyclic peptide are as follows:

selecting sheep blood cells, centrifuging at 4 ℃ at 3000 r/min for 10min, discarding the supernatant, washing the lower layer red blood cells with normal saline for 3 times, and then resuspending into 3% red blood cell suspension. Add 100uL of peptide solutions at different concentrations to EP tubes, the final concentration of peptide solution in each tube (ug/mL) was: 256. 128, 64, 32, 16, 4, 100uL of red blood cell suspension was added. Each set is provided with three parallel sets. The negative control group was added with an equal volume of physiological saline, and the positive control group was added with 100uL of 0.1% Triton-X100. And (3) culturing the reaction solution in a biochemical incubator at 37 ℃ for 0.5h, taking out, centrifuging at 3000 r/min for 10min, absorbing 100uL of supernatant, transferring to a 96-well plate, and measuring the absorbance at the wavelength of 570nm by using an enzyme-labeling instrument. The experiment was repeated three times and the data averaged.

Hemolysis rate [ (OD)_{Test well}-OD_{Negative hole})/(OD_{Positive hole}-OD_{Negative hole})]×100％

The results are shown in Table 3.

TABLE 3 results of the determination of hemolytic activity of antibacterial cyclic peptide

A lower value of hemolysis of the antimicrobial peptide indicates a lower toxicity of the antimicrobial peptide. As can be seen from the table, in comparison with the control

Compared with the peptide, the hemolytic toxicity of the antibacterial cyclic peptide CPeptide-A, CPeptide-B, CPeptide-C, CPeptide-D, CPeptide-F is smaller, while the hemolytic toxicity of the antibacterial cyclic peptide CPeptide-E is larger at high concentrations of 256 and 128, but the antibacterial cyclic peptide CPeptide-E still has certain research value based on good antibacterial activity.

The present invention is not limited to the above-described embodiments, and those skilled in the art will understand that: various changes, modifications, substitutions and alterations can be made to the embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the claims and their equivalents.

Sequence listing

<110> Chongqing university of science and technology

<120> anti-multidrug-resistant cyclopeptide and application thereof

<130>

<160>6

<210> 1

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<213> Artificial Synthesis

<220>

<223>CPeptide-A

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Arg-Arg-Trp-Trp-Trp-Arg

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<213> Artificial Synthesis

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<223>CPeptide-B

<400> 2

Arg-Trp-Arg-Trp-Arg-Trp

1 6

<210> 3

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<212> PRT

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<223>CPeptide-C

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Arg-Arg-Trp-Trp-Trp-Arg-Trp

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<223>CPeptide-D

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Arg-Arg-Trp-Trp-Arg-Arg-Trp

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<212> PRT

<213> Artificial Synthesis

<220>

<223>CPeptide-E

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Arg-Arg-Trp-Trp-Trp-Trp-Arg-Arg

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<213> Artificial Synthesis

<220>

<223>CPeptide-F

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Arg-Arg-Trp-Trp-Trp-Trp-Arg-Arg

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Claims

1. The cyclic peptide for resisting multiple drug resistance bacteria is characterized in that the amino acid sequence is any one of the following six types:

CPeptide-A：R-R-W-W-W-R(Arg-Arg-Trp-Trp-Trp-Arg)

CPeptide-B:R-W-R-W-R-W(Arg-Trp-Arg-Trp-Arg-Trp)

CPeptide-C:R-R-W-W-W-R-R(Arg-Arg-Trp-Trp-Trp-Arg-Trp)

CPeptide-D:R-R-W-W-R-R-W(Arg-Arg-Trp-Trp-Arg-Arg-Trp)

CPeptide-E:R-R-W-W-W-W-R-R(Arg-Arg-Trp-Trp-Trp-Trp-Arg-Arg)

CPeptide-F:R-R-W-W-R-R-R-W(Arg-Arg-Trp-Trp-Arg-Arg-Arg-Trp)

the structural formulas after cyclization are respectively as follows:

CPeptide-A:

CPeptide-B:

CPeptide-C:

CPeptide-D:

CPeptide-E:

CPeptide-F

。

2. the use of the anti-multidrug resistance cyclopeptide according to claim 1 for antimicrobial applications.

3. The use of the multidrug-resistant cyclopeptide according to claim 1 in the resistance to multidrug-resistant acinetobacter baumannii, escherichia coli, and staphylococcus aureus.