CN110054664B - Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof - Google Patents

Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof Download PDF

Info

Publication number
CN110054664B
CN110054664B CN201910320083.5A CN201910320083A CN110054664B CN 110054664 B CN110054664 B CN 110054664B CN 201910320083 A CN201910320083 A CN 201910320083A CN 110054664 B CN110054664 B CN 110054664B
Authority
CN
China
Prior art keywords
fmoc
ano
resin
leu
side chain
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN201910320083.5A
Other languages
Chinese (zh)
Other versions
CN110054664A (en
Inventor
倪京满
王锐
钟超
王一杰
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Lanzhou University
Original Assignee
Individual
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Individual filed Critical Individual
Priority to CN201910320083.5A priority Critical patent/CN110054664B/en
Publication of CN110054664A publication Critical patent/CN110054664A/en
Application granted granted Critical
Publication of CN110054664B publication Critical patent/CN110054664B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/08—Peptides having 5 to 11 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/542—Carboxylic acids, e.g. a fatty acid or an amino acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Organic Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Epidemiology (AREA)
  • Immunology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Genetics & Genomics (AREA)
  • Molecular Biology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Communicable Diseases (AREA)
  • Oncology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Peptides Or Proteins (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

The invention designs and synthesizes a side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid, which takes linear amphipathic alpha-helical natural antibacterial peptide Anoplin as a template, combines the strategy of enhancing enzymolysis stability by D-type amino acid replacement and enhancing antibacterial activity by fatty acid modification of a side chain of the replaced D-type amino acid, and obtains a side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D-type amino acid with a brand new structure n And Ano-D4,7-7C n N=4-16. The in-vitro bacteriostasis experiment, the PI staining flow cytometry experiment and the enzymolysis stability experiment show that the side chain fatty acid modified antibacterial peptide analogue containing the D-type amino acid has strong drug-resistant bacteria resistance and high enzymolysis stability; compared with the traditional antibiotics, the novel antibacterial peptide analogue has good application prospect in the aspect of development of clinical antibacterial medicines.

Description

Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof
Technical Field
The invention relates to the technical field of biochemistry, relates to a D-type amino acid-containing side chain fatty acid modified antibacterial peptide analogue with a brand new structure, and synthesis and application thereof, and in particular relates to a D-type amino acid-containing side chain fatty acid modified antibacterial peptide analogue with strong drug-resistant bacteria activity and high enzymolysis stability, and synthesis and application thereof.
Background
In recent years, due to the abuse of traditional antibiotics in clinical medicine, the emergence of resistant strains, i.e. "superbacteria", which exhibit resistance to most or all available antibiotics (Health Econ.1996May-Jun;5 (3): 217-26). Polymyxin B and polymyxin E (also known as colistin) are widely used as cationic peptide antibiotics in the 60 s of the 20 th century, but their clinical application is greatly reduced in the 70 s of the 20 th century due to serious toxicity problems. Although the use of these two antibacterial agents has been resuscitated as the 20 th century 70 epidemic gram-negative bacteria became more resistant, the last antibiotic, unfortunately, polymyxin-resistant "superbacteria" also appeared successively (Expert Rev Anti Infect Ther.2012Aug;10 (8): 917-34;Biomed Res Int.2015;2015:679109). Needless to say, the development of a new class of antibiotics has become critical (Lancet effect Dis.2013Dec;13 (12): 1057-98). As a novel antibiotic having a great potential, antibacterial peptides (AMPs), particularly cationic antibacterial peptides, have received great attention because of their broad-spectrum antibacterial activity and rapid sterilization (Chembiochem. 2015Jan 19;16 (2): 242-53). AMPs are typically produced by a variety of biological organisms including bacteria, fungi, plants, insects, amphibians, crustaceans, fish and mammals (Clin Microbiol rev.2006jul;19 (3): 491-511). Most importantly, bacteria are less likely to develop resistance to antimicrobial peptides that do not have specific targets of action than traditional antibiotics. The mode of action of the antibacterial peptide generally involves non-specific interactions with the bacterial cytoplasmic membrane, allowing the antibacterial peptide in the resulting bacterial membrane to accumulate, increasing the permeability of the membrane and losing barrier function, ultimately leading to leakage of the bacterial content and death (Eur. J. Biochem.2001,268,5589-5600;Nat Rev Microbiol.2005Mar;3 (3): 238-50).
However, AMPs, while being able to combat "superbacteria" as compared to traditional antibiotics, are limited in their clinical use as ideal antibacterial agents, poor antibacterial activity, toxicity to host cells, intolerance to physiological conditions, susceptibility to enzymatic degradation, and high manufacturing costs due to complex designs. A large number of researches show that the introduction of D-type amino acid can effectively avoid the degradation of protease and improve the enzymolysis stability of the antibacterial peptide (Sci Rep.2017Jul 31;7 (1): 6953;Chem Biol Drug Des.2006Feb;67 (2): 162-73), but the introduction of D-type amino acid generally leads to the reduction of the antibacterial activity thereof. The fatty acid is used as an important component of phospholipid of biological cell membrane, has higher hydrophobicity, is introduced into the antibacterial peptide, and is beneficial to enhancing the affinity of the antibacterial peptide to bacterial cell membrane by increasing the hydrophobicity of the antibacterial peptide, so that the antibacterial activity of the antibacterial peptide is enhanced, and the fatty acid can also reduce the degradation of protease, enhance the enzymolysis stability of the antibacterial peptide and prolong the in vivo acting time of the antibacterial peptide (Biochem J,2005, 385 (Pt 1): 135-43;Biophys Chem,2015, 199: 25-33).
Disclosure of Invention
One of the objects of the present invention is: provides a side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid with a brand new structure, strong antibacterial activity and high enzymolysis stability.
The second object of the present invention is: provides the application of the antibacterial peptide analogue in the development of clinical antibacterial medicaments.
The third object of the present invention is: provides a method for synthesizing the antibacterial peptide analogue.
Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid
The invention relates to a side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid, which is characterized in that 4-position or 7-position of a part of D-type amino acid substitution analogue Ano-D4,7 of parent peptide Anoplin is introduced with D-type special unnatural amino acid Fmoc-D-Lys (Mtt) -OH with side chain protecting group Mtt, then the side chain protecting group is removed, and the side chain of the D-type special unnatural amino acid is subjected to fatty acid (C) n N=4-16) to obtain side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D type amino acid with novel structure n ，Ano-D4,7-7C n ，n＝4-16。
The structural formula is shown as follows:
Gly-Leu-Leu-D-Lys(C n )-Arg-Ile-D-Lys-Thr-Leu-Leu-NH2
wherein n=4-16, designated Ano-D4,7-4C n ；
Gly-Leu-Leu-D-Lys-Arg-Ile-D-Lys(C n )-Thr-Leu-Leu-NH2
Wherein n=4-16, designated Ano-D4,7-7C n ；
The invention relates to a synthesis method of a side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid, which comprises the following process steps:
1、Ano-D4,7-4C n is synthesized by (a)
The Fmoc-Leu-OH, HOBT, HBTU, DIEA is dissolved and mixed uniformly in DMF, and the Fmoc-Leu-resin is subjected to condensation reaction with MBHA resin from which Fmoc protecting groups are removed, so that Fmoc-Leu-resin is obtained; the amino acids Fmoc-Leu-OH, fmoc-Thr (tBu) -OH, fmoc-D-Lys (Boc) -OH, fmoc-Ile-OH, fmoc-Arg (pbf) -OH, fmoc-D-Lys (Mtt) -OH, fmoc-Leu-OH and Fmoc-Gly-OH are condensed in sequence by the same method to obtain Fmoc-Gly-Leu-Leu-D-Lys (Mtt) -Arg-Ile-D-Lys-Thr-Leu-Leu-resin, namely Fmoc-Ano-D4,7-4 (Mtt) -resin;
removing side chain Mtt protecting group of Fmoc-Ano-D4,7-4 (Mtt) -resin obtained above with DCM solution containing 1% TFA by volume fraction to obtain Ano-D4,7-4 (NH) 2 ) -resin; dissolving fatty acid, HOBT, HBTU and DIEA in DMF, mixing with Ano-D4,7-4 (NH 2 ) Condensation reaction of the resin to obtain Ano-D4,7-4C n -resin; ano-D4,7-4C n Cutting and purifying the resin to obtain the analogue Ano-D4,7-4C of the antibacterial peptide n ，n＝4-16。
2、Ano-D4,7-7C n Is synthesized by (a)
The Fmoc-Leu-OH, HOBT, HBTU, DIEA is dissolved and mixed uniformly in DMF, and the Fmoc-Leu-resin is subjected to condensation reaction with MBHA resin from which Fmoc protecting groups are removed, so that Fmoc-Leu-resin is obtained; the amino acids Fmoc-Leu-OH, fmoc-Thr (tBu) -OH, fmoc-D-Lys (Mtt) -OH, fmoc-Ile-OH, fmoc-Arg (pbf) -OH, fmoc-D-Lys (Boc) -OH, fmoc-Leu-OH and Fmoc-Gly-OH are condensed in sequence by the same method to obtain Fmoc-Gly-Leu-Leu-D-Lys-Arg-Ile-D-Lys (Mtt) -Thr-Leu-Leu-resin, namely Fmoc-Ano-D4,7-7 (Mtt) resin;
removing side chain Mtt protecting group of Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above with DCM solution containing 1% TFA by volume fraction to obtain Ano-D4,7-7 (NH) 2 )-resin; dissolving fatty acid, HOBT, HBTU and DIEA in DMF, mixing with Ano-D4,7-7 (NH 2 ) Condensation reaction of the resin to obtain Ano-D4,7-7C n -resin; ano-D4,7-7C n Cutting and purifying the resin to obtain the analogue Ano-D4,7-7C of the antibacterial peptide n ，n＝4-16。
The concentrations of the amino acids, the fatty acids, the HOBT, the HBTU and the DIEA in DMF are respectively 20-100mg/mL,20-100mg/mL,10-40mg/mL,20-100mg/mL and 20-60mg/mL; the molar mass ratio of each amino acid, fatty acid, HOBT, HBTU and the MBHA resin with Fmoc protecting groups removed is 6:1-3:1, and the molar mass ratio of DIEA to the MBHA resin with Fmoc protecting groups removed is 6:1-12:1.
The cleavage reagent is a mixed solution of TFA, triisopropylsilane and water in a volume ratio of 9.5:0.25:0.25.
The purification process comprises the steps of freeze-drying crude peptide to obtain freeze-dried powder, and then performing RP-HPLC separation; RP-HPLC purification conditions were mobile phase A: aqueous 0.05% tfa, mobile phase B: acetonitrile solution of 0.05% tfa; and (3) carrying out linear gradient elution, and collecting effluent liquid of a main absorption peak.
The mass spectrum identification shows that the method successfully synthesizes the side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D-type amino acid with novel structure n And Ano-D4,7-7C n ，n＝4-16。
(II) in vitro Activity study of side chain fatty acid modified antibacterial peptide analog containing D-type amino acid
1. Bacteriostasis experiment
The minimum inhibitory concentration, i.e., the MIC value, of the above-described antimicrobial peptide analogs was determined using a classical double dilution method. The experimental strains selected include standard normal strains: coli ATCC 25922,P.aeruginos ATCC 27853,K.pneumoniae ATCC 700603,S.aureus ATCC 25923,B.subtilis ATCC 23857,S.epidermidis ATCC 12228 and clinically isolated multi-drug resistant strain: baumannii 9828,A.baumannii 9840,P.aeruginosa 1240,P.aeruginosa 1190,E.coli 8500,E.coli 8040,S.aureus 4800,S.aureus 5200. The specific experimental method is as follows: experimental bacteria grown overnight in MH medium to log phase were diluted to1×10 6 CFU/mL bacterial suspension; dissolving the antibacterial peptide analogue in sterile water, preparing a series of peptide solutions with different concentrations of 1-128 mu mol/L by a double dilution method, mixing with the bacterial suspension in equal volume, incubating for 18-24h at 37 ℃ in a 96-well culture plate, and observing that the minimum concentration without obvious bacterial growth is the minimum inhibitory concentration MIC; the antibiotics Erythromycin, kanamycin and Penicillin are used as positive control medicines; the above experiment was repeated three times in parallel, and the results are shown in tables 1 and 2.
TABLE 1 minimum inhibitory concentration against Standard Normal Strain
Figure BDA0002034362060000041
/>
Figure BDA0002034362060000051
TABLE 2 minimum inhibitory concentration against multidrug resistant strains
Figure BDA0002034362060000052
Table 1 shows that the side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D amino acid n And Ano-D4,7-7C n The antibacterial activity of the strain is obviously improved compared with that of parent peptide Anoplin, and the antibacterial activity of the strain is partially better than that of the traditional antibiotics; table 2 shows that the side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D amino acid n And Ano-D4,7-7C n The strain has stronger antibacterial activity to clinically separated multi-drug resistant bacteria strains, and compared with parent peptide Anoplin, the antibacterial activity of the strain is obviously improved and is superior to that of the traditional antibiotics; with increasing fatty acid chain length, antibacterial activity of the antibacterial peptide analogue against both standard strains and multi-drug resistant bacteria is significantly enhanced, but when fatty acid chain length is increased to a certain extent, antibacterial activity is no longer increased.
2. Flow cytometry experiments
PI staining flow cytometry was performed using a standard strain of escherichia coli (ATCC 25922) to detect bacterial membrane disruption by the antimicrobial peptide analogs. The specific experimental method is as follows: e.coli cultured to log phase was diluted to 10X 10 8 CFU/mL, washed with PBS (10 mm, ph 7.4) and resuspended in half volume to give bacterial suspension; the antibacterial peptide analogue was dissolved in PBS at a concentration of 8 XMIC, mixed with the same volume of the above bacterial suspension, incubated at 37℃for 2 hours, stained with Pyridine Iodide (PI) in the dark for 15min, washed off the excess dye with PBS, and examined for uptake of PI fluorescence by flow cytometry, and the results are shown in FIG. 9.
FIG. 9 shows that the side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D-type amino acid n And Ano-D4,7-7C n All have better bacterial cell membrane destructive power; as the length of the side chain fatty acid increases, the ability of the antibacterial peptide analogue to destroy bacterial cell membranes increases significantly, which can be explained by the significant increase in antibacterial activity described above.
3. Enzymolysis stability test
The peptide solution was incubated with trypsin solutions of different concentrations (1 mg/mL,0.5mg/mL,0.2mg/mL,0.1 mg/mL) at 37℃for 1h and 6h; after 15min of inactivation at 60 ℃, the minimum inhibitory concentration MIC for e.coli ATCC 25922 in different trypsin solutions was determined as described above with respect to the minimum inhibitory concentration assay, and the results are shown in table 3.
TABLE 3 minimum inhibitory concentration of E.coli ATCC 25922 under different trypsin concentrations
Figure BDA0002034362060000061
Control: minimum inhibitory concentration on E.coli ATCC 25922 in the absence of trypsin
The results in Table 3 show that the parent peptide Anoplin loses antibacterial activity under trypsin environments with different concentrations and shows low stability; side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D-type amino acid n And Ano-D4,7-7C n In trypsin environment with different concentrationsThe antimicrobial activity is not lost, the MIC is only slightly changed, and the antimicrobial activity is better, so that the side chain fatty acid modified antimicrobial peptide analogue containing D-type amino acid has higher stability in trypsin environment, and the stability is obviously better than that of parent peptide.
In conclusion, the invention combines the strategy of enhancing enzymolysis stability by replacing D-type amino acid and enhancing the activity of resisting drug-resistant bacteria by modifying the side chain of the replaced D-type amino acid by taking the linear amphipathic alpha-helical natural antibacterial peptide Anoplin as a template, so as to obtain the side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid with a brand new structure. The in vitro biological activity research results show that the side chain fatty acid modified antibacterial peptide analogue containing the D-type amino acid has strong antibacterial activity against drug-resistant bacteria and high enzymolysis stability, and has good application prospect in clinical antibacterial drug development.
Drawings
FIG. 1 shows the antibacterial peptide analogues Ano-D4,7-4C of the invention 4 Mass spectrum of (3);
FIG. 2 shows the antibacterial peptide analogues Ano-D4,7-4C of the invention 8 Mass spectrum of (3);
FIG. 3 shows the antibacterial peptide analogues Ano-D4,7-4C of the invention 12 Mass spectrum of (3);
FIG. 4 shows the antibacterial peptide analogues Ano-D4,7-4C of the invention 16 Mass spectrum of (3);
FIG. 5 shows the antibacterial peptide analogues Ano-D4,7-7C of the invention 4 Mass spectrum of (3);
FIG. 6 shows the antibacterial peptide analogues Ano-D4,7-7C of the invention 8 Mass spectrum of (3);
FIG. 7 shows the antibacterial peptide analogues Ano-D4,7-7C of the invention 12 Mass spectrum of (3);
FIG. 8 shows the antibacterial peptide analogues Ano-D4,7-7C of the invention 16 Mass spectrum of (3);
FIG. 9 is a graph showing the results of PI-stained flow cytometry experiments with the antimicrobial peptide analogs of the present invention; in the figure, the control group, anoplin group, ano-D4,7-4C are sequentially arranged from left to right and from top to bottom 8 Group Ano-D4,7-4C 12 Group Ano-D4,7-7C 8 Group and Ano-D4,7-7C 12 A group.
Detailed Description
The synthesis of the D-amino acid-containing side chain fatty acid-modified antibacterial peptide analog of the present invention is further described below by way of specific examples.
Example 1: ano-D4,7-4C 4 Is synthesized by (a)
(1) Resin activation and pretreatment
Accurately weighing 0.47g of MBHA resin (0.43 mmol/g), placing in a polypeptide solid-phase synthesizer, swelling with DCM solution for 30min, and checking by ninhydrin chromogenic method to obtain colorless transparent resin, which indicates that the resin is normal.
(2) Synthesis of Fmoc-Ano-D4,7-4 (Mtt) -resin
Removing Fmoc protecting groups from the MBHA resin with normal detection through DMF solution containing 20% of piperidine by volume fraction, and detecting the resin by an ninhydrin chromogenic method to show that the protecting groups are removed; fmoc-Leu-OH (212 mg), HOBT (81 mg), HBTU (228 mg) and DIEA (0.2 mL) are dissolved and uniformly mixed in 5-10mL of DMF, added into a synthesizer, mixed with the MBHA resin without the Fmoc protecting group, and subjected to condensation reaction for 1h; the ninhydrin color-rendering method is used for testing, and the resin is colorless and transparent, so that the condensation reaction is successful, and Fmoc-Leu-resin is obtained; the method is the same as above, and the following amino acids are subjected to condensation reaction in turn: fmoc-Leu-OH (212 mg), fmoc-Thr (tBu) -OH (239 mg), fmoc-D-Lys (Boc) -OH (281 mg), fmoc-Ile-OH (212 mg), fmoc-Arg (pbf) -OH (390 mg), fmoc-D-Lys (Mtt) -OH (376 mg), fmoc-Leu-OH (212 mg), fmoc-Gly-OH (238 mg), HOBT, HBTU and DIEA were used in the same amounts, wherein the Fmoc-D-Lys (Mtt) -OH condensation reaction time was 1.5h, the balance was 1h, giving Fmoc-Gly-Leu-D-Lys (Mtt) -Arg-Ile-D-Lys-Thr-Leu-resin, i.e., fmoc-An-D4, 7-4 (Mtt) -in;
(3)Ano-D4,7-4C 4 synthesis of resin
Removing side chain Mtt protecting group of Fmoc-Ano-D4,7-4 (Mtt) -resin with DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4 (NH) 2 ) -resin; separately, butyric anhydride (C) n N=4; 0.87 mL), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mLDissolving in DMF, mixing, adding into synthesizer, and removing side chain Mtt protecting group from Ano-D4,7-4 (NH) 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-4C 4 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4C 4 -resin。
(4) Polypeptide cleavage
Ano-D4,7-4C 4 Cutting the resin by taking a mixed solution of TFA, triisopropylsilane and water with the water volume ratio of 9.5:0.25:0.25 as a cutting reagent, extracting by using glacial ethyl ether and water, and freeze-drying to obtain crude peptide freeze-dried powder;
(5) Polypeptide purification
Separating and purifying the crude peptide lyophilized powder obtained by freeze drying by RP-HPLC, collecting effluent, freeze drying, and identifying by mass spectrum to obtain Ano-D4,7-4C 4 The molecular weight is 1223Da, and the mass spectrum is shown in figure 1; wherein, RP-HPLC purification conditions: mobile phase a 0.05% tfa/water; mobile phase B0.05% tfa/acetonitrile; and (3) carrying out linear gradient elution, and collecting effluent liquid of a main absorption peak.
Example 2: ano-D4,7-4C 8 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-4 (Mtt) -resin
As in example 1.
(3)Ano-D4,7-4C 8 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-4 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4 (NH) 2 ) -resin; separately octanoic anhydride (C) n N=8; 1.53 mL), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) were dissolved in 5-10mL DMF and mixed, and added to a synthesizer together with the aforementioned Ano-D4,7-4 (NH) with the side chain Mtt protecting group removed 2 ) Resin mixing, condensation reaction1.5 hours; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-4C 8 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4C 8 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-4C was obtained by mass spectrometry 8 The molecular weight is 1279Da, and the mass spectrum is shown in figure 2.
Example 3: ano-D4,7-4C 12 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-resin
As in example 1.
(3)Ano-D4,7-4C 12 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-4 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4 (NH) 2 ) -resin; separately adding dodecanoic acid (C) n N=12; 240 mg), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mL DMF was dissolved and mixed evenly, and added to a synthesizer together with Ano-D4,7-4 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-4C 12 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4C 12 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, A was obtained by mass spectrometryno-D4,7-4C 12 The molecular weight is 1335Da, and the mass spectrum is shown in figure 3.
Example 4: ano-D4,7-4C 16 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-resin
As in example 1.
(3)Ano-D4,7-4C 16 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-4 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4 (NH) 2 ) -resin; separately separating hexadecanoic acid (C) n N=16; 307 mg), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mL DMF was dissolved and mixed, and added to a synthesizer together with Ano-D4,7-4 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-4C 16 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-4C 16 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-4C was obtained by mass spectrometry 16 The molecular weight is 1391Da, and the mass spectrum is shown in FIG. 4.
Example 5: ano-D4,7-7C 4 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-7 (Mtt) -resin
Removing Fmoc protecting groups from the MBHA resin with normal detection through DMF solution containing 20% of piperidine by volume fraction, and detecting the resin by an ninhydrin chromogenic method to show that the protecting groups are removed; fmoc-Leu-OH (212 mg), HOBT (81 mg), HBTU (228 mg) and DIEA (0.2 mL) are dissolved and uniformly mixed in 5-10mL of DMF, added into a synthesizer, mixed with the MBHA resin without the Fmoc protecting group, and subjected to condensation reaction for 1h; the ninhydrin color-rendering method is used for testing, and the resin is colorless and transparent, so that the condensation reaction is successful, and Fmoc-Leu-resin is obtained; the method is the same as above, and the following amino acids are subjected to condensation reaction in turn: fmoc-Leu-OH (212 mg), fmoc-Thr (tBu) -OH (239 mg), fmoc-D-Lys (Mtt) -OH (376 mg), fmoc-Ile-OH (212 mg), fmoc-Arg (pbf) -OH (390 mg), fmoc-D-Lys (Boc) -OH (281 mg), fmoc-Leu-OH (212 mg), fmoc-Gly-OH (238 mg), HOBT, HBTU and DIEA were used in the same amounts, wherein the Fmoc-D-Lys (Mtt) -OH condensation reaction time was 1.5h, the balance was 1h, to obtain Fmoc-Gly-Leu-D-Lys-Arg-Ile-D-Lys (Mtt) -Thr-Leu-Leu-resin, namely Fmoc-An-D4, 7-4 (Mtt) -in;
(3)Ano-D4,7-7C 4 synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7 (NH) 2 ) -resin; separately, butyric anhydride (C) n N=4; 0.87 mL), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mL DMF was dissolved and mixed, and added to a synthesizer together with the aforementioned Ano-D4,7-7 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-7C 4 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7C 4 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-7C was identified by mass spectrometry 4 The molecular weight is 1223Da, and the mass spectrum is shown in figure 5.
Example 6: ano-D4,7-7C 8 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-7 (Mtt) -resin
Same as in example 5.
(3)Ano-D4,7-7C 8 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7 (NH) 2 ) -resin; separately octanoic anhydride (C) n N=8; 1.53 mL), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) were dissolved in 5-10mL DMF and mixed, and added to a synthesizer together with the aforementioned Ano-D4,7-7 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-7C 8 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7C 8 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-7C was identified by mass spectrometry 8 The molecular weight is 1279Da, and the mass spectrum is shown in figure 6.
Example 7: ano-D4,7-7C 12 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-7 (Mtt) -resin
Same as in example 5.
(3)Ano-D4,7-7C 12 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7 (NH) 2 )-resin; separately adding dodecanoic acid (C) n N=12; 240 mg), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mL DMF was dissolved and mixed evenly, and added to a synthesizer together with Ano-D4,7-7 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-7C 12 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7C 12 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-7C was identified by mass spectrometry 12 The molecular weight is 1335Da, and the mass spectrum is shown in figure 7.
Example 8: ano-D4,7-7C 16 Is synthesized by (a)
(1) Resin activation and pretreatment
As in example 1.
(2) Synthesis of Fmoc-Ano-D4,7-7 (Mtt) -resin
Same as in example 5.
(3)Ano-D4,7-7C 16 Synthesis of resin
Removing side chain Mtt protecting group from Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above by DCM solution containing 1% TFA by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7 (NH) 2 ) -resin; separately separating hexadecanoic acid (C) n N=16; 307 mg), HOBT (81 mg), HBTU (228 mg), DIEA (0.2 mL) in 5-10mL DMF was dissolved and mixed, and added to a synthesizer together with Ano-D4,7-7 (NH) with the side chain Mtt protecting group removed 2 ) -resin mixing, condensation reaction for 1.5h; the ninhydrin color-development method is used for testing, and the resin is colorless and transparent, which indicates that the condensation reaction is complete, thus obtaining Ano-D4,7-7C 16 -resin; removing N-terminal Fmoc protecting group with DMF solution containing 20% piperidine by volume fraction, and detecting by ninhydrin chromogenic method to obtain Ano-D4,7-7C 16 -resin。
(4) Polypeptide cleavage
As in example 1.
(5) Polypeptide purification
As in example 1, ano-D4,7-7C was identified by mass spectrometry 16 The molecular weight is 1335Da, and the mass spectrum is shown in figure 8.

Claims (9)

1. The side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid is characterized in that the structural formula of the antibacterial peptide analogue is as follows:
Gly-Leu-Leu-D-Lys(C n )-Arg-Ile-D-Lys-Thr-Leu-Leu-NH 2
Ano-D4,7-4C n ，C n is octanoic acid, dodecanoic acid or hexadecanoic acid;
or (b)
Gly-Leu-Leu-D-Lys-Arg-Ile-D-Lys(C n )-Thr-Leu-Leu-NH 2
Ano-D4,7-7C n ，C n Is octanoic acid, dodecanoic acid or hexadecanoic acid.
2. The use of a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid as claimed in claim 1 in the preparation of a clinical antibacterial medicament.
3. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to claim 1, wherein the method comprises the following steps:
introducing a part of D-type amino acid substitution analogue Ano-D4,7 of Ano-D4,7 sequence of the Anoplin into the D-type special unnatural amino acid Fmoc-D-Lys (Mtt) -OH with a side chain protecting group Mtt by adopting a classical solid phase synthesis method to synthesize Ano-D4,7-4 (Mtt) -resin or Ano-D4,7-7 (Mtt) -resin; then removing side chain protecting groups, and modifying the side chains of the D-type special unnatural amino acid by fatty acids with different lengths; finally, the side chain fatty acid modified antibacterial peptide analogue Ano-D4,7-4C containing D-type amino acid is obtained after cutting and purifying n Or Ano-D4,7-7C n Wherein C n Is octanoic acid, dodecanoic acid or hexadecanoic acid.
4. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to claim 3, wherein the method for synthesizing the antibacterial peptide analogue specifically comprises the following steps:
(1)Ano-D4,7-4C n is synthesized by (a)
The Fmoc-Leu-OH, HOBT, HBTU, DIEA is dissolved and mixed uniformly in DMF, and the Fmoc-Leu-resin is subjected to condensation reaction with MBHA resin from which Fmoc protecting groups are removed, so that Fmoc-Leu-resin is obtained; the amino acids Fmoc-Leu-OH, fmoc-Thr (tBu) -OH, fmoc-D-Lys (Boc) -OH, fmoc-Ile-OH, fmoc-Arg (pbf) -OH, fmoc-D-Lys (Mtt) -OH, fmoc-Leu-OH and Fmoc-Gly-OH are condensed in sequence by the same method to obtain Fmoc-Gly-Leu-Leu-D-Lys (Mtt) -Arg-Ile-D-Lys-Thr-Leu-Leu-resin, namely Fmoc-Ano-D4,7-4 (Mtt) -resin;
removing the side chain Mtt protecting group of Fmoc-Ano-D4,7-4 (Mtt) -resin obtained above with DCM solution containing 1% TFA by volume fraction to obtain Fmoc-Ano-D4,7-4 (NH) 2 ) -resin; the fatty acid, HOBT, HBTU and DIEA were dissolved in DMF and mixed with Fmoc-Ano-D4,7-4 (NH 2 ) Condensation reaction of the resin to obtain Fmoc-Ano-D4,7-4C n -resin; fmoc-protecting group is removed from Fmoc-Ano-D4,7-4Cn-resin, and then the Fmoc-protecting group is cut and purified to obtain the antibacterial peptide analogue Ano-D4,7-4C n ，C n Is octanoic acid, dodecanoic acid or hexadecanoic acid;
(2)Ano-D4,7-7C n is synthesized by (a)
The Fmoc-Leu-OH, HOBT, HBTU, DIEA is dissolved and mixed uniformly in DMF, and the Fmoc-Leu-resin is subjected to condensation reaction with MBHA resin from which Fmoc protecting groups are removed, so that Fmoc-Leu-resin is obtained; the amino acids Fmoc-Leu-OH, fmoc-Thr (tBu) -OH, fmoc-D-Lys (Mtt) -OH, fmoc-Ile-OH, fmoc-Arg (pbf) -OH, fmoc-D-Lys (Boc) -OH, fmoc-Leu-OH and Fmoc-Gly-OH are condensed in sequence by the same method to obtain Fmoc-Gly-Leu-Leu-D-Lys-Arg-Ile-D-Lys (Mtt) -Thr-Leu-Leu-resin, namely Fmoc-Ano-D4,7-7 (Mtt) -resin;
removing the side chain Mtt protecting group from the Fmoc-Ano-D4,7-7 (Mtt) -resin obtained above with DCM solution containing 1% TFA by volume fraction to obtain Fmoc-Ano-D4,7-7 (NH) 2 ) -resin; respectively separating fatty acid and HOBT, HBTU and DIEA were dissolved in DMF and mixed with Fmoc-Ano-D4,7-7 (NH 2 ) Condensation reaction of the resin to obtain Fmoc-Ano-D4,7-7C n -resin; fmoc-Ano-D4,7-7C n -cutting and purifying after Fmoc-protecting group is removed from resin to obtain antibiotic peptide analogue Ano-D4,7-7C n ，C n Is octanoic acid, dodecanoic acid or hexadecanoic acid.
5. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to claim 4, wherein the concentration of each amino acid, each fatty acid, HOBT, HBTU and DIEA in DMF is 20-100mg/mL,20-100mg/mL,10-40mg/mL,20-100mg/mL and 20-60mg/mL, respectively.
6. The method for synthesizing a D-amino acid-containing side chain fatty acid-modified antibacterial peptide analogue according to claim 4 or 5, wherein the molar mass ratio of each amino acid, each fatty acid, HOBT, HBTU and the Fmoc-removed MBHA resin is 6:1 to 3:1, and the molar mass ratio of diea to the Fmoc-removed MBHA resin is 6:1 to 12:1.
7. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to any one of claims 3 to 5, wherein the cleavage reagent is a mixed solution of TFA, triisopropylsilane and water in a volume ratio of 9.5:0.25:0.25.
8. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to any one of claims 3 to 5, wherein the purification process comprises the steps of freeze drying and then performing RP-HPLC separation and purification; RP-HPLC purification conditions were mobile phase A: aqueous 0.05% tfa, mobile phase B: acetonitrile solution of 0.05% tfa; and (3) carrying out linear gradient elution, and collecting effluent liquid of a main absorption peak.
9. The method for synthesizing a side chain fatty acid modified antibacterial peptide analogue containing a D-type amino acid according to claim 6, wherein the purification process comprises freeze-drying the obtained crude peptide, and then separating and purifying by RP-HPLC; RP-HPLC purification conditions were mobile phase A: aqueous 0.05% tfa, mobile phase B: acetonitrile solution of 0.05% tfa; and (3) carrying out linear gradient elution, and collecting effluent liquid of a main absorption peak.
CN201910320083.5A 2019-04-19 2019-04-19 Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof Active CN110054664B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201910320083.5A CN110054664B (en) 2019-04-19 2019-04-19 Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201910320083.5A CN110054664B (en) 2019-04-19 2019-04-19 Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof

Publications (2)

Publication Number Publication Date
CN110054664A CN110054664A (en) 2019-07-26
CN110054664B true CN110054664B (en) 2023-05-26

Family

ID=67319804

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201910320083.5A Active CN110054664B (en) 2019-04-19 2019-04-19 Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof

Country Status (1)

Country Link
CN (1) CN110054664B (en)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110563802B (en) * 2019-09-04 2023-04-07 倪京满 Group of antibacterial peptide analogues containing N-methylated amino acid and N-terminal fatty acid modification, and synthetic method and application thereof
CN113072619B (en) * 2021-04-09 2023-05-30 倪京满 Alpha helix antibacterial short peptide with high antibacterial activity and low toxicity and application thereof

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106632600A (en) * 2016-10-19 2017-05-10 倪京满 D-type non-natural amino acid containing antimicrobial peptide analog, synthesis therefor and application of D-type non-natural amino acid containing antimicrobial peptide analog
CN109265518A (en) * 2018-10-10 2019-01-25 倪京满 N- terminal aliphatic acid modified antimicrobial peptide analogues and its synthesis and application with high enzymatic hydrolysis stability and strong antibacterial activity

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106632600A (en) * 2016-10-19 2017-05-10 倪京满 D-type non-natural amino acid containing antimicrobial peptide analog, synthesis therefor and application of D-type non-natural amino acid containing antimicrobial peptide analog
CN109265518A (en) * 2018-10-10 2019-01-25 倪京满 N- terminal aliphatic acid modified antimicrobial peptide analogues and its synthesis and application with high enzymatic hydrolysis stability and strong antibacterial activity

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
Àngel Oliveras et al.Antimicrobial activity of linear lipopeptides derived from BP100 towards plant pathogens.《PLOS ONE》.2018, *
Synthesis and biological activity of lipophilic analogs of the cationic antimicrobial active peptide anoplin;Kostas Chionis et al;《Journal of peptide science》;20161231;摘要，第3页右栏第1段-第4页左栏第1段，第4页右栏第2段-第5页左栏最后1段，表2，图4 *

Also Published As

Publication number Publication date
CN110054664A (en) 2019-07-26

Similar Documents

Publication Publication Date Title
Čeřovský et al. Lucifensin, a novel insect defensin of medicinal maggots: synthesis and structural study
CN111363010B (en) Symmetrical short-sequence antibacterial peptide analogue and application thereof
CN110054664B (en) Side chain fatty acid modified antibacterial peptide analogue containing D-type amino acid and synthesis and application thereof
CN113754784A (en) Cell-penetrating antibacterial peptide and application thereof
EP3122763A1 (en) Antimicrobial peptide dendrimers
CN116874614B (en) Antibacterial polypeptide APH171 with high activity and low cracking effect, and preparation method and application thereof
CN111057134A (en) Antibacterial peptides with broad-spectrum antibacterial activity, low toxicity and no induced drug resistance and application thereof
CN107344958B (en) Antibacterial pentapeptide derivative and application thereof
CN106749559B (en) Antibacterial peptide based on cell-penetrating peptide Tat (49-57) and synthesis method thereof
CN110938112A (en) Fatty acid modified ultrashort-sequence antibacterial peptide analogue and application thereof
KR100879211B1 (en) Novel antimicrobial peptide and its application derived from echiuroid worm, Urechis unicinctus
CN112778401B (en) Caprylic acid acylation modified antibacterial peptide and application thereof
CN105837675B (en) Cationic antibacterial peptides and preparation method thereof
CN110437308B (en) Beta amino acid-containing antibacterial peptide analogue with specific activity on pseudomonas aeruginosa and application thereof
CN107266546B (en) Antibacterial peptide HlDFS1 and application thereof
CN113999297B (en) Antibacterial peptide hrNCM and preparation method and application thereof
CN111410681B (en) Antibacterial peptide analogue with small repeat unit sequence and application thereof
CN108409833B (en) Novel candida-killing polypeptide FCP1 and preparation method thereof
CN113698464A (en) Antibacterial peptide HeHamp II-4 (63-88) and application thereof
CN113527458A (en) Antibacterial peptide HeHamp II-2 (63-86) and application thereof
CN113735956A (en) Antibacterial peptide CCM7WC, and preparation method and application thereof
CN112625106A (en) Antibacterial polypeptide compound, synthesis method and application thereof
KR20020049443A (en) Novel antibiotic peptide derived from ribosomal protein l1 of helicobacter pylori and use thereof
CN115010788B (en) N-terminal fatty acid modified low-toxicity broad-spectrum antibacterial peptide analogue with antibacterial film activity and application thereof
CN113583090B (en) Arnopyrin modified peptide with antibacterial activity and synthesis method and application thereof

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant
TR01 Transfer of patent right

Effective date of registration: 20230919

Address after: 730000 No. 222 Tianshui South Road, Chengguan District, Gansu, Lanzhou

Patentee after: LANZHOU University

Address before: 730000 room 609, 680 Dingxi Road, Chengguan District, Lanzhou City, Gansu Province

Patentee before: Ni Jingman

Patentee before: Wang Rui

TR01 Transfer of patent right