CN116813712A - Antibacterial peptide W33 with alpha-helical structure and rich in Trp, and preparation method and application thereof - Google Patents
Antibacterial peptide W33 with alpha-helical structure and rich in Trp, and preparation method and application thereof Download PDFInfo
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- CN116813712A CN116813712A CN202310714576.3A CN202310714576A CN116813712A CN 116813712 A CN116813712 A CN 116813712A CN 202310714576 A CN202310714576 A CN 202310714576A CN 116813712 A CN116813712 A CN 116813712A
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- 239000003910 polypeptide antibiotic agent Substances 0.000 title claims abstract description 66
- 238000002360 preparation method Methods 0.000 title abstract description 8
- 230000000844 anti-bacterial effect Effects 0.000 claims abstract description 30
- 239000003814 drug Substances 0.000 claims abstract description 5
- 230000003252 repetitive effect Effects 0.000 claims abstract description 4
- 208000022362 bacterial infectious disease Diseases 0.000 claims abstract 2
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 27
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 26
- 102000044503 Antimicrobial Peptides Human genes 0.000 claims description 18
- 108700042778 Antimicrobial Peptides Proteins 0.000 claims description 18
- 229920001184 polypeptide Polymers 0.000 claims description 12
- 230000015572 biosynthetic process Effects 0.000 claims description 8
- 238000001514 detection method Methods 0.000 claims description 8
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 7
- 238000003786 synthesis reaction Methods 0.000 claims description 7
- 230000002949 hemolytic effect Effects 0.000 claims description 6
- 230000003993 interaction Effects 0.000 claims description 5
- 238000000034 method Methods 0.000 claims description 5
- 238000001819 mass spectrum Methods 0.000 claims description 4
- 150000007523 nucleic acids Chemical group 0.000 claims description 4
- 238000004007 reversed phase HPLC Methods 0.000 claims description 3
- NQIHMZLGCZNZBN-PXNSSMCTSA-N Trp-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@H](CC=3C4=CC=CC=C4NC=3)N)C(O)=O)=CNC2=C1 NQIHMZLGCZNZBN-PXNSSMCTSA-N 0.000 claims description 2
- 230000009435 amidation Effects 0.000 claims description 2
- 238000007112 amidation reaction Methods 0.000 claims description 2
- 239000003674 animal food additive Substances 0.000 claims description 2
- 230000003013 cytotoxicity Effects 0.000 claims description 2
- 231100000135 cytotoxicity Toxicity 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims description 2
- 239000007790 solid phase Substances 0.000 claims description 2
- 108010045269 tryptophyltryptophan Proteins 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 claims 1
- 231100001274 therapeutic index Toxicity 0.000 abstract description 12
- 241000894006 Bacteria Species 0.000 abstract description 8
- 241000192125 Firmicutes Species 0.000 abstract description 5
- 208000015181 infectious disease Diseases 0.000 abstract description 2
- 229940079593 drug Drugs 0.000 abstract 1
- 210000003743 erythrocyte Anatomy 0.000 description 10
- 210000004027 cell Anatomy 0.000 description 5
- 230000001580 bacterial effect Effects 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 239000012071 phase Substances 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 206010018910 Haemolysis Diseases 0.000 description 3
- 239000003085 diluting agent Substances 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 230000008588 hemolysis Effects 0.000 description 3
- 230000002401 inhibitory effect Effects 0.000 description 3
- 239000013641 positive control Substances 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- 230000000845 anti-microbial effect Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 125000003345 AMP group Chemical group 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 241000194032 Enterococcus faecalis Species 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 1
- 241000293869 Salmonella enterica subsp. enterica serovar Typhimurium Species 0.000 description 1
- 241000191967 Staphylococcus aureus Species 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 210000004899 c-terminal region Anatomy 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- 229940032049 enterococcus faecalis Drugs 0.000 description 1
- 229920006227 ethylene-grafted-maleic anhydride Polymers 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 210000005007 innate immune system Anatomy 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000005142 microbroth dilution method Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000010532 solid phase synthesis reaction Methods 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- GPRLSGONYQIRFK-MNYXATJNSA-N triton Chemical compound [3H+] GPRLSGONYQIRFK-MNYXATJNSA-N 0.000 description 1
Classifications
-
- 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
- C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/142—Amino acids; Derivatives thereof
- A23K20/147—Polymeric derivatives, e.g. peptides or proteins
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
-
- 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
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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- Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Polymers & Plastics (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Oncology (AREA)
- Animal Behavior & Ethology (AREA)
- Genetics & Genomics (AREA)
- Communicable Diseases (AREA)
- Biophysics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Molecular Biology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Biochemistry (AREA)
- Animal Husbandry (AREA)
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- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention provides an antibacterial peptide W33 with an alpha-helical structure and rich in Trp, and a preparation method and application thereof. The antibacterial peptide W33 is shown as SEQ ID No.1, and the preparation method comprises the following steps: based on a standard alpha helix heptapeptide repetitive sequence 'abcdefg', leu is selected to be placed at the a and d positions to form a Leu zipper structure, trp is added at the c and f positions, arg is added at the rest positions, and the heptapeptide sequence is repeated for 2 times to reach enough positive charge number, so that the antibacterial peptide W33 is obtained. The application of the antibacterial peptide in medicines for treating gram-negative bacteria and/or gram-positive bacteria infectious diseases. The antibacterial peptide W33 has a therapeutic index of 98.84, has good cell selectivity, has potential to become a broad-spectrum antibacterial medicament for treating gram-positive bacteria and gram-negative bacteria infection, and has high application value.
Description
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a preparation method and application of an antibacterial peptide W33 with an alpha-helical structure and rich in Trp.
Background
Antibacterial peptide (Antimicrobial peptides, AMPs) is an important component of the innate immune system of organisms as a small peptide substance with broad-spectrum antibacterial activity. Compared with the traditional antibiotics, the antibacterial peptide has the advantages of wide sources, broad-spectrum antibacterial property, difficult generation of drug resistance, safety, no residue and quick action, and the advantages make the antibacterial peptide an ideal antibiotic substitute. However, natural antibacterial peptides have the limitations of low activity, poor stability, short half-life and great difficulty in synthesis in animal husbandry, and researchers aim at brand-new designs of antibacterial peptides in order to solve the problems. The stable secondary structure is the precondition of the interaction between the antibacterial peptide and bacterial cell membrane, thus ensuring the good antibacterial activity of the antibacterial peptide. Alpha-helices have been widely studied in recent years as the most common structure in antimicrobial peptides. The existing natural antibacterial peptide has the problems of low activity, large synthesis difficulty caused by overlong amino acid sequence and the like.
Disclosure of Invention
Based on the defects, the invention aims to provide a preparation method of an antibacterial peptide W33 with an alpha-helical structure and rich in Trp, which solves the problems of low activity and high synthesis difficulty caused by overlong amino acid sequences of the existing natural antibacterial peptide.
The technical scheme adopted by the invention is as follows: an antibacterial peptide W33 with alpha-helical structure and rich Trp has an amino acid sequence shown in SEQ ID No.1 and a C-terminal of-NH 2 Amidation.
Further, the molecular formula is shown as formula (I),
the invention also aims to provide a preparation method of the antibacterial peptide W33 with the alpha-helical structure and rich in Trp, which comprises the following steps:
(1) Firstly, using an alpha-helix heptapeptide repetitive sequence abcdefg as a basis, selecting Leu to be placed at an a position and a d position to form a Leu zipper structure, and facilitating the formation of an alpha-helix structure; adding Trp at c and f positions to ensure that the interaction of Trp-Trp improves the stability of an alpha-helix structure, thereby improving antibacterial activity, adding Arg at other positions to increase the number of positive charges, repeating a heptapeptide sequence for 2 times to reach the sufficient number of positive charges, and designing a polypeptide, wherein the amino acid sequence of the polypeptide is shown as SEQ ID No. 1;
(2) The designed polypeptide is synthesized by adopting a solid-phase chemical synthesis method, and is further subjected to reversed-phase high-performance liquid chromatography purification and mass spectrum identification to prepare the polypeptide, and the polypeptide is subjected to antibacterial activity detection, cytotoxicity detection and hemolytic activity detection, and finally named as antibacterial peptide W33.
Another object of the present invention is to provide the use of an antibacterial peptide W33 of alpha-helical structure enriched in Trp as described above for the preparation of a medicament for the treatment of infectious diseases of gram-positive bacteria or/and gram-negative bacteria.
It is another object of the present invention to provide a nucleic acid fragment encoding the antibacterial peptide W33 as described above.
It is another object of the present invention to provide a recombinant vector into which the nucleic acid fragment as described above is inserted.
Another object of the present invention is to provide a feed additive comprising the above-mentioned antimicrobial peptide W33.
The invention has the following advantages and beneficial effects: the invention can achieve good antibacterial effect by using only 14 amino acids, effectively reduces the synthesis cost, and simultaneously inhibits the toxicity of the antibacterial peptide. The antibacterial peptide W33 obtained is subjected to antibacterial and hemolytic activity detection, and the antibacterial peptide W33 is found to have strong antibacterial activity on gram-positive bacteria and gram-negative bacteria of a plurality of detection such as escherichia coli, pseudomonas aeruginosa, salmonella typhimurium, staphylococcus aureus, enterococcus faecalis and the like, and has a therapeutic index of 98.84 and good cell selectivity. In conclusion, the antibacterial peptide W33 with the alpha-helical structure and rich in Trp has potential to be a broad-spectrum antibacterial medicament for treating gram-positive bacteria and gram-negative bacteria infection, and has high application value.
Drawings
FIG. 1 is a reversed-phase high performance liquid chromatogram of antibacterial peptide W23;
FIG. 2 is a reversed-phase high performance liquid chromatogram of antibacterial peptide W33;
FIG. 3 is a mass spectrum of the antibacterial peptide W23;
FIG. 4 is a mass spectrum of the antibacterial peptide W33;
FIG. 5 is a three-dimensional structure prediction diagram of antibacterial peptide W23;
FIG. 6 is a three-dimensional structure prediction diagram of antibacterial peptide W33;
FIG. 7 is a spiral wheel projection of the antimicrobial peptide W23;
FIG. 8 is a spiral wheel projection of the antimicrobial peptide W33;
Detailed Description
The present invention will be described in further detail with reference to the following examples and drawings.
Example 1
Design of antibacterial peptides
By using an alpha-helix heptapeptide repetitive sequence 'abcdefg' as a basis, leu is selected to be placed at an a position and a d position to form a Leu zipper structure, so that the formation of the alpha-helix structure is facilitated; when the paired Trps are added at the positions b and e, the interaction of the paired Trps can stabilize the alpha-helical structure, the antibacterial activity is obviously improved, arg is added at the other positions to increase the positive charge number of the antibacterial peptide, and the heptapeptide sequence is repeated for 2 times to reach the sufficient positive charge number, so that the antibacterial peptide W23 is obtained; when the paired Trps are added at the positions c and f, the interaction of the paired Trps can stabilize the alpha-helical structure, the antibacterial activity is obviously improved, arg is added at the other positions to increase the positive charge number of the antibacterial peptide, and the heptapeptide sequence is repeated for 2 times to reach the sufficient positive charge number, so that the antibacterial peptide W33 is obtained. The sequence and physicochemical parameters of the antimicrobial peptide are shown in Table 1.
TABLE 1 antibacterial peptide W23, W33 sequences and physicochemical parameters
a Average hydrophobicity was calculated by HeliQuest analysis;
b the relative moment of hydrophobicity was calculated by HeliQuest analysis.
Example 2
Synthesis and identification of antibacterial peptides
The designed antibacterial peptide is synthesized by a polypeptide synthesizer through a solid phase synthesis method, and is identified by an electrospray mass spectrometry method and purified by reverse-phase high performance liquid chromatography to obtain a target compound for subsequent structure and activity determination.
Reverse phase high performance liquid chromatograms of the antibacterial peptides W23 and W33 with alpha-helical structures and rich in Trp are shown in figures 1 and 2.
The mass spectrograms of the antibacterial peptides W23 and W33 with alpha-helical structures and rich in Trp are shown in figures 3 and 4.
Example 3
Computer software predicts the secondary structure of antibacterial peptides
The three-dimensional structure of the antibacterial peptide is predicted by using on-line structure prediction software I-TASSER and inputting an amino acid sequence. The three-dimensional structure prediction diagrams of the antibacterial peptides W23 and W33 with alpha-helical structures rich in Trp are shown in figures 5 and 6. From the figure, it can be seen that Trp-rich α -helix antibacterial peptides W23, W33 exhibit a standard α -helix structure. Indicating that the antibacterial peptide with the alpha-helical structure rich in Trp is successfully designed. Spiral-wheel projection diagrams of Trp-rich α -helical antibacterial peptides W23, W33 predicted by the HeliQuest program are shown in fig. 7-8.
Example 4
Antibacterial activity of antibacterial peptide
Antibacterial peptide antibacterial activity is known by measuring Minimum Inhibitory Concentration (MIC) of the antibacterial peptide. The designed and successfully synthesized antimicrobial peptides were formulated as 2.56mM stock solutions for bioactivity assays. The strain to be tested was inoculated into fresh MHB broth medium, placed in shaking table at 37℃and shaking speed of 220rpm overnight for culturing, and transferred to new, sterile MHB for culturing the next day to logarithmic phase of cell growth. OD of bacteria Using a Spectrophotometer 600nm The value is regulated to about 0.4, and the bacterial liquid is diluted 1000 times for standby. The MIC of the antimicrobial peptide was determined by a micro broth dilution method using a sterile 96-well plate with 0.01% acetic acid (0.2% BSA) as a diluent. The antibacterial peptide stock was added to the dilution of the 96-well plate, subjected to a 2-fold gradient dilution, and then 50. Mu.L of bacterial suspension was added to each well. Wherein bacterial-containing MHB was used as a positive control, and non-sterile MHB medium was used as a negative control. Incubating the culture plate in a 37 ℃ incubator for 18-20h, and observing the minimum antimicrobial peptide concentration without microorganism growth with naked eyes to obtain the MIC of the determined polypeptide to the detected bacteria. Three independent replicates were performed, two replicates each. The minimum inhibitory concentration of the antimicrobial peptides is shown in table 2.
TABLE 2 antibacterial Activity of antibacterial peptides W23, W33 (. Mu.M)
As can be seen from the table, the antimicrobial peptides W23 and W33 all showed extremely strong antimicrobial activity against the 8 strains tested, and the MIC values thereof were all in the range of 2-8. Mu.M, wherein the antimicrobial peptide W33 showed a stronger antimicrobial effect.
Example 5
Hemolytic Activity of antibacterial peptides
To assess the safety of the antimicrobial peptides, the haemolytic behaviour of peptides on human blood erythrocytes (hRBC) in the concentration range of 1-128. Mu.M was studied.
Fresh blood of healthy persons was collected, 1mL was centrifuged at 1000 Xg for 5min, the supernatant was discarded, and erythrocytes were collected. The collected erythrocytes were washed 3 times with PBS buffer and finally resuspended with 10mL of PBS. PBS was added to erythrocytes in a ratio such that the positive control value was around 1. Taking a sterile 96-well culture plate, taking PBS buffer solution as a diluent, adding the antibacterial peptide storage solution into the diluent of the 96-well culture plate, carrying out gradient multiple ratio dilution by 2 times, adding 50 mu L of red blood cell suspension into each well, uniformly mixing with 50 mu L of antibacterial peptide solution with different concentrations, and incubating for 1h at a constant temperature in a 37 ℃ incubator. Centrifuging at 4deg.C and 1000 Xg for 10min; 50. Mu.L of each well was centrifuged to obtain a supernatant,transfer to a new 96-well plate and absorbance at 570nm was measured using a microplate reader. Wherein 50. Mu.L of red blood cells plus 50. Mu.L of 0.1% Triton x-100 was used as a positive control and 50. Mu.L of red blood cells plus 50. Mu.L of PBS buffer was used as a negative control. By measuring the minimum concentration (HC) of the antibacterial peptide at 50% hemolysis of human red blood cells caused by W23 and W33 50 ) The biocompatibility of both was evaluated, and the cell selectivity of both was further evaluated by calculating the W23, W33 Therapeutic Index (TI), see table 3.
TABLE 3 biocompatibility of antibacterial peptides W23, W33
a Geometric Mean (GM) of antimicrobial peptide pair for determining minimum inhibitory concentration of bacteria;
b HC 50 is the lowest concentration of antimicrobial peptide that causes 50% hemolysis of human blood erythrocytes (hRBC), 256 μm was used to calculate the therapeutic index when no detectable hemolytic activity was observed at 128 μm;
c the Therapeutic Index (TI) is HC 50 Ratio to GM.
By measuring the minimum concentration (HC) of the antimicrobial peptides W23, W33 at 50% hemolysis of human blood erythrocytes (hRBC) 50 ) HC found in W33 50 Value [(s) ]>128 μm) far higher HC than W23 50 The value (16. Mu.M) indicates that the biosafety of W33 is high. Further calculation of the W23, W33 Therapeutic Index (TI) assessed the cell selectivity of both, higher TI values indicating that the antimicrobial peptide possesses higher cell selectivity. Wherein the TI value (98.84) of the antibacterial peptide W33 is significantly higher than the TI value (5.19) of the antibacterial peptide W23, which indicates that the antibacterial peptide W33 has better cell selectivity.
Claims (7)
1. An antibacterial peptide W33 with alpha-helical structure and rich Trp is characterized in that the amino acid sequence is shown in SEQ ID No.1, and the C end adopts-NH 2 Amidation.
2. An antibacterial peptide W33 with an alpha-helical structure rich in Trp according to claim 1 and having a molecular formula shown in formula (I),
3. the method for preparing the antibacterial peptide W33 with the alpha-helical structure rich in Trp according to claim 1, comprising the following steps:
(1) Firstly, using an alpha-helix heptapeptide repetitive sequence abcdefg as a basis, selecting Leu to be placed at an a position and a d position to form a Leu zipper structure, and facilitating the formation of an alpha-helix structure; adding Trp at c and f positions to ensure that the interaction of Trp-Trp improves the stability of an alpha-helix structure, thereby improving antibacterial activity, adding Arg at other positions to increase the number of positive charges, repeating a heptapeptide sequence for 2 times to reach the sufficient number of positive charges, and designing a polypeptide, wherein the amino acid sequence of the polypeptide is shown as SEQ ID No. 1;
(2) The designed polypeptide is synthesized by adopting a solid-phase chemical synthesis method, and is further subjected to reversed-phase high-performance liquid chromatography purification and mass spectrum identification to prepare the polypeptide, and the polypeptide is subjected to antibacterial activity detection, cytotoxicity detection and hemolytic activity detection, and finally named as antibacterial peptide W33.
4. Use of an antibacterial peptide W33 of Trp-rich α -helix structure according to claim 1 for the manufacture of a medicament for the treatment of a gram positive or/and gram negative bacterial infectious disease.
5. A nucleic acid fragment encoding the antimicrobial peptide W33 of claim 1.
6. A recombinant vector, wherein the nucleic acid fragment according to claim 5 is inserted into the recombinant vector.
7. A feed additive comprising the antibacterial peptide W33 of claim 1.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117285599A (en) * | 2023-10-11 | 2023-12-26 | 东北农业大学 | Cell penetrating antibacterial peptide 5VT resisting intracellular bacteria, and preparation method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20070117746A1 (en) * | 2003-12-17 | 2007-05-24 | A12 Limited | Treatment of viral infections |
| CN108570103A (en) * | 2018-04-03 | 2018-09-25 | 东北农业大学 | One kind is rich in tryptophan antibacterial peptide WK12 and its preparation method and application |
| CN114014915A (en) * | 2021-10-13 | 2022-02-08 | 东北农业大学 | Broad-spectrum antibacterial alpha helical peptide and preparation method and application thereof |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20070117746A1 (en) * | 2003-12-17 | 2007-05-24 | A12 Limited | Treatment of viral infections |
| CN108570103A (en) * | 2018-04-03 | 2018-09-25 | 东北农业大学 | One kind is rich in tryptophan antibacterial peptide WK12 and its preparation method and application |
| CN114014915A (en) * | 2021-10-13 | 2022-02-08 | 东北农业大学 | Broad-spectrum antibacterial alpha helical peptide and preparation method and application thereof |
Non-Patent Citations (4)
| Title |
|---|
| QING-QUAN MA 等: ""The effects of Leu or Val residues on cell selectivity of α-helical peptides"", 《PROTEIN PEPT LETT》, vol. 18, no. 11, pages 1112 * |
| YINFENG LV 等: ""Antimicrobial properties and membrane-active mechanism of a potential α-helical antimicrobial derived from cathelicidin PMAP-36"", 《PLOS ONE》, vol. 9, no. 1, pages 10 * |
| 单安山 等: ""抗菌肽的功能、研发与应用"", 《中国农业科学》, vol. 45, no. 11, pages 2249 - 2259 * |
| 来振衡 等: ""抗菌肽在动物生产中应用的研究进展"", 《动物营养学报》, vol. 34, no. 6, pages 3401 - 3410 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117285599A (en) * | 2023-10-11 | 2023-12-26 | 东北农业大学 | Cell penetrating antibacterial peptide 5VT resisting intracellular bacteria, and preparation method and application thereof |
| CN117285599B (en) * | 2023-10-11 | 2024-04-09 | 东北农业大学 | Cell penetrating antibacterial peptide 5VT resisting intracellular bacteria, and preparation method and application thereof |
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