KR102019153B1 - Antimicrobial peptide derived from rock bream and uses thereof - Google Patents

Abstract

The present invention provides an antimicrobial peptide derived from Oplegnathus fasciatus showing high antimicrobial activity against various marker strains and excellent salt and thermal stability.

Description

Antimicrobial peptide derived from rock bream and uses approximately

The present invention relates to an antibacterial peptide derived from dolme, more particularly to an antibacterial peptide derived from dolme and to its use.

Antibiotics have been used in humans for over a hundred years, freeing people from bacterial diseases and increasing productivity of livestock and seafood. However, due to the recent antibiotic residues and accumulation of antibiotic resistant strains, their use is very limited, and antibiotics used in livestock and aquatic products are completely prohibited or only partially allowed. Therefore, the development of substances that can replace these antibiotics is required, and various alternative materials research for this is being carried out. Among various alternatives, antimicrobial peptides are the natural antibiotics produced by living organisms and there is no danger of resistant strains.They have the broadest antimicrobial activity against various microorganisms and are easy to manufacture through high-synthesis or microbial cultivation process. .

 Antimicrobial peptides (AMPs) are antimicrobial activities of up to about 100 short-length peptides first discovered in the epidermal secretions of frogs in the 1960s. Positive charges such as lysine, arginine, and histidine Contains a large number of amino acids. Therefore, by attaching to the negatively-charged microbial cell membrane to destroy the microbial cell membrane has an antimicrobial activity. Almost all living things are known to produce from microorganisms to plants, invertebrates, vertebrates and humans, and have unique antimicrobial properties against a wide range of microorganisms, including bacteria, fungi, viruses and parasites. Antimicrobial peptides have been shown to be the most important defenses in the innateve immune system because they act as a defense against various external invading microorganisms in a short time.In addition to these antimicrobial activities, they have antioxidant, anti-inflammatory, anti-insect and wound healing activities. It has been reported to have a multifunctional property. In addition, there is a low possibility of emergence of resistant strains due to physical antibacterial action mechanism and has the advantage of easy synthesis due to the small size. Therefore, the antimicrobial peptides, which are natural bioprotective substances, are recently expected to be used in various ways, and are particularly attracting attention in the field of antibiotic replacement, various additives, and pharmaceutical development. In this regard, the Republic of Korea Patent No. 1660258 discloses a new antimicrobial peptide derived from the sea bream and its use.

However, in the case of the prior art, it is difficult to expect a uniform effect due to factors such as potential toxicity and not showing sufficient antimicrobial activity because the yield is not high when the peptide is separated.

The present invention is to solve various problems, including the above problems, it is an object of the present invention to provide an antimicrobial peptide derived from dolmedo exhibiting excellent antimicrobial activity against various indicator strains. However, these problems are exemplary, and the scope of the present invention is not limited thereby.

According to one aspect of the invention, there is provided an antimicrobial peptide comprising the amino acid sequence set forth in SEQ ID NO: 3 (RFLKLMMKLL).

According to another aspect of the invention, there is provided an antimicrobial composition containing an antimicrobial peptide as an active ingredient.

According to another aspect of the invention, there is provided a feed additive containing an antimicrobial peptide as an active ingredient.

According to another aspect of the present invention, there is provided a composition for controlling an anti-scotch caterpillar, which contains an antimicrobial peptide as an active ingredient.

According to another aspect of the invention, there is provided an anticancer composition containing an antimicrobial peptide as an active ingredient.

According to one embodiment of the present invention made as described above, it is possible to implement the effect of producing a sea bream-derived antimicrobial peptide showing high antimicrobial activity against various strains and stable to heat and salt. Of course, the scope of the present invention is not limited by these effects.

1 is a schematic diagram of the secondary structure and RB-LBP-II-5A amino acid substitution for the production of RB-LBP-II-5A antibacterial peptide derived from Doldo LBP of the present invention.
Figure 2 is a photograph analyzing the antimicrobial activity of the natural form of the RB-LBP-II-5N antimicrobial peptide against the indicator strain.
Figure 3a is a photograph analyzing the antimicrobial activity of the RB-LBP-II-5A antimicrobial peptide against the indicator strain using the URDA method.
Figure 3b is a photograph analyzing the antimicrobial activity of the RB-LBP-II-5A antibacterial peptide against the indicator strain using the URDA method.
Figure 4 is a photograph analyzing the stability of the antimicrobial activity against heat and salt of the RB-LBP-II-5A antibacterial peptide.
5 is a graph analyzing the antiparasitic activity to evaluate the multifunctionality of the RB-LBP-II-5A antibacterial peptide.
6 is a graph analyzing the anticancer activity of the RB-LBP-II-5A antibacterial peptide.

Definition of Terms:

As used herein, the term "AMPs (Antimicrobial peptides)" is an antimicrobial peptide, the following properties are known. Many organisms on Earth have an immune system that can protect themselves from the environment they are in, and are a versatile bioactive substance of primary defense means that produce them to protect themselves against pathogenic microbes. It plays an important role and is reported to be present in almost all organisms on earth, from mammals to insects and plants as a natural antibiotic (Zasloff, Nature 415: 389-395, 2002). Antimicrobial peptides were first detected in the epidermal secretions of frogs ( Bombina variegate ) in 1962 (Kiss and Michl, Toxicon 1: 33-39,1962; Monatshefte f ㆌ r Chemie -Chemical Monthly, 101: 182-189.1970), 1971 Various antimicrobial peptides have been identified from various organisms such as bee venom, and many studies have been conducted on their properties and functions. So far, the most valid hypothesis of the mechanism of action of antimicrobial peptides is the Shai-Matsuzaki-Huang (SMH) model.The peptide synthesized in the body has a hydrophobic and hydrophilic moiety with a suitable structure and a positively charged hydrophilic moiety. Binding to the cell membrane of negatively charged bacteria (Matsuzaki, Biochimica et Biophysica Acta 1462: 1-10. 1999; Shai, Biochimica et Biophysica Acta, 1462: 55-70. 1999; Huang, Biochemistry 39: 8347-8352. 2000 Yang et al., Biophysical Journal, 79: 2002-2009. 2000). At this time, the binding force between the peptide and the cell membrane is known to be about 2-3 times stronger than general Ca ²⁺ , Mg ²⁺ . The hydrophobic portion of the peptide bound to the cell membrane then interacts with the hydrophobic portion of the cell membrane phospholipids to form pores that change the permeability of the cell membrane and eventually destroy the cell (Christensen et al., Proceeding of the National Academy of Sciences). 85: 0.5072 to 5076, 1988).

As used herein, the term “lipopolysaccharide binding protein (LBP)” is a glycoprotein with a molecular weight of 60,000 present in serum and biosynthesized in large quantities in the liver during acute phase reactions. LBP binds to the lipid A portion of Gram-negative bacteria LPS (lipid polysaccharide), opsonizes erythrocytes encoded by Gram-negative bacteria or LPS, and activates macrophages by LPS by more than 1,000-fold.

Detailed description of the invention:

According to one aspect of the invention, there is provided an antimicrobial peptide comprising the amino acid sequence set forth in SEQ ID NO: 3 (RFLKLMMKLL).

In the antimicrobial peptide, the C-terminus may be amidated.

According to another aspect of the invention, there is provided an antimicrobial composition containing an antimicrobial peptide as an active ingredient.

In the antimicrobial composition, it may have an antimicrobial activity against gram negative bacteria, gram positive bacteria or fungi.

In the antimicrobial composition, the Gram-positive bacteria are B. cereus, B. subtilis, S. mutans, S. iniae, S. aureus, L. garvieae, S. parauberis, S. aureus 0203 , S. aureus 0204 , S. aureus . pyogenes 0206, or S. aureus 3795 and the Gram-negative bacteria are E. coli, P. aeruginosa, V. alginoalyticus, V. anguillarum, V. parahaemolyticus, V. vulnificus, V. harveyi, K. pneumoniae, E. tzarda , P. stuartii, E. coli ML35p, P. aeruginosa 0225, E. coli 0238, S. typhimurium 0240, K. oxytoca 0248, K. aerogenes 0249, E. cloacae 0252, or E. coli 1A814 and the fungus May be C. albicans .

According to another aspect of the invention, there is provided a feed additive containing an antimicrobial peptide as an active ingredient.

According to another aspect of the present invention, there is provided a composition for controlling an anti-scotch caterpillar, which contains an antimicrobial peptide as an active ingredient.

According to another aspect of the invention, there is provided an anticancer composition containing an antimicrobial peptide as an active ingredient.

In the anticancer composition, the cancer may be uterine cancer or lung cancer.

The effective amount of the antimicrobial peptide of the present invention may vary depending on the type of affected part, the site of application, the number of treatments, the treatment time, the dosage form, the condition of the patient, the type of adjuvant, and the like. The amount used is not particularly limited, but may be 0.01 μg / kg / day to 10 mg / kg / day. The daily dose may be administered once or in two or three times a day at appropriate intervals or may be administered intermittently at intervals of several days.

The antimicrobial peptide of the present invention may be contained in an amount of 0.1-100% by weight based on the total weight of the composition, and may further include appropriate carriers, excipients and diluents commonly used in the preparation of pharmaceutical compositions. In addition, the preparation of the pharmaceutical composition may be used as additives for the preparation of solid or liquid. The additive for preparation may be either organic or inorganic. Examples of excipients include lactose, sucrose, white sugar, glucose, cornstarch, starch, talc, sorbet, crystalline cellulose, dextrin, kaolin, calcium carbonate and silicon dioxide. As the binder, for example, polyvinyl alcohol, polyvinyl ether, ethyl cellulose, methyl cellulose, gum arabic, tragacanth, gelatin, shellac, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, calcium citrate, Dextrin, pectin, and the like. Examples of the lubricant include magnesium stearate, talc, polyethylene glycol, silica, hardened vegetable oil, and the like. As a coloring agent, if it is normally permitted to add to a pharmaceutical, all can be used. These tablets and granules can be appropriately coated according to sugar, gelatin coating and other needs. Moreover, preservatives, antioxidants, etc. can be added as needed.

The antimicrobial peptides of the present invention may be prepared in any formulation conventionally prepared in the art (e.g., Remington's Pharmaceutical Science, New Edition; Mack Publishing Company, Easton PA), and the form of the formulation is not particularly limited. These formulations are described in Remington's Pharmaceutical Science, 15 ^th Edition, 1975, Mack Publishing Company, Easton, Pennsylvania 18042 (Chapter 87: Blaug, Seymour), a prescription generally known for all pharmaceutical chemistries.

The antimicrobial peptide of the present invention can be administered orally or parenterally, and preferably, by parenteral administration, intravenous injection, subcutaneous injection, intracerebroventricular injection, intracerebrospinal fluid injection, intramuscular injection. And intraperitoneal infusion.

The inventors of the present invention suggest that bio-specific antimicrobial peptide research based on antimicrobial activity requires various analytical processes such as pure separation of target substance in each process, confirmation of antimicrobial activity of separated fractions, post-translational modification and terminal sequence analysis, and pure separation. As a result of careful efforts to solve the existing analytical methods and the difficulty of sequencing analysis, we conducted a sequence-based antimicrobial peptide study using genome analysis information to secure various antimicrobial peptide candidates and showed excellent antimicrobial activity The present invention was completed by analyzing the excellent antimicrobial activity and properties of LBP protein-derived antimicrobial peptides.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, the present invention is not limited to the embodiments disclosed below, but can be implemented in various forms, and the following embodiments are intended to complete the disclosure of the present invention, to those skilled in the art It is provided to inform you completely.

Example 1 Genome Analysis

According to an embodiment of the present invention, a sequence-based antimicrobial peptide study using genome analysis information was performed to secure various antimicrobial peptide candidates. Oplegnathus fasciatus genome analysis results based on the post-translational protein sequence based on the antibacterial peptide characteristics helix secondary structure, 2 to 50 or less short sequence, including a large number of positively charged amino acids, both hydrophobicity and hydrophilicity As a result of analyzing the short protein sequences satisfying the amphipathic, low Bowman index, etc., some short peptide sequences were obtained as antimicrobial peptide candidates. Among the nucleotide sequence-based amino acid sequences in the obtained genomic data, 20 or more various antimicrobial peptide candidate sequences were obtained, and among them, peptide properties of about 1.3 kDa Doldom lipopolysaccharide binding protein (LBP) sequences were between 2 and 50 amino acids. As a result of analyzing the number, helix structure, charge value, Bowman index, etc. and the most suitable sequence for antimicrobial peptide characteristics, 16 amino acid sequence (SEQ ID NO: 1) of PQIAQRFPGLMMKLLV among the peptide candidates was determined. It was confirmed that it had a value that best matched the characteristic (see Table 1). Information on the antimicrobial peptide candidates derived from the dolme LBP gene is shown in Table 1 below.

Peptide Name Sequence (SEQ ID NO) M.W. p.I. Pure charge Hydrophobicity (kcal / mol) Bowman index (kcal / mol) SEQ ID NO: RB-LBP-II-5 PQIAQRFPGLMMKLLV 1842.33 11.01 +2  0.53 -0.16 One RB-LBP-II-5N RFPGLMMKLL 1205.4 11.00 +2 -1.01 -0.29 2 RB-LBP-II-5A RFLKLMMKLL 1292.75 11.17 +3 -1.04 -0.13 3

Example 2: Antimicrobial Peptide Preparation

After analyzing the helix wheel, which is a protein secondary structure using the sequence of Table 1, some amino acid residues of the N-terminus and C-terminus were removed. A peptide (RFPGLMMKLL, SEQ ID NO: 2) consisting of 10 amino acids from which N-terminal five amino acids were removed from 16 amino acid sequences in Doldom LBP protein was constructed and named RB-LBP-II-5N (Table 1). ). The designed peptide was obtained by requesting solid phase synthesis from Peptron, Inc., a peptide synthesis company. In addition, to analyze the secondary structure of the 10 amino acid sequence to prepare the antimicrobial peptide with increased antimicrobial activity, the sequence of the front and rear amino acids in the helix structure and the structure of the amino acids and residues of the top and bottom amino acids were analyzed. Amphiphilicity was assessed to specifically replace the third amino acid residue with other amino acid residues that matched the properties of the antimicrobial peptide. Specifically, the third amino acid (proline) and the fourth amino acid (glycine) of the RB-LBP-II-5N peptide, respectively, were leucine. And a peptide substituted with lysine and named it RB-LBP-II-5A (SEQ ID NO: 3) (Table 1 and FIG. 1). In addition, RFLKLMMKLL-NH ₂ peptide was designed in which the carboxyl group (C-term) was amidated with NH ₂ to increase the structural stability and antibacterial activity of the RB-LBP-IU-5A peptide. , Inc.) to obtain a solid phase synthesis.

Example 3: Antimicrobial Activity Determination Assay (URDA)

One fungus by dissolving the RB-LBP-II-5N and RB-LBP-II-5A antimicrobial peptides secured according to an embodiment of the present invention in a 0.01% acetic acid solution at a concentration of 1,000-31.3 ㎍ / ml, Determination of antimicrobial activity against 35 pathogenic microorganisms, including 15 Gram-positive bacteria, 19 Gram-negative bacteria, and antibiotic-resistant strains, was performed.

First, using the optimal growth medium of each indicator strain, then incubated for 18 hours at an appropriate culture temperature and diluted with the same medium so that the bacteria 10 ⁸ CFU / ml or yeast 10 ⁶ CFU / ml corresponding to OD ₆₀₀ = 0.1. Then, 0.5 ml of the strain culture diluted in 9.5 ml of an under-lay gel containing 0.01% culture medium, 1% agarose and 10 mM phosphate buffer (pH 6.5) was added to each plate, followed by mixing. The plate was solidified by dispensing. After a hole of 2.2 mm in diameter in the hard under-lay gel, the peptide dissolved in 10 μl of 0.01% acetic acid was injected. In addition, 10 ml of over-lay gel containing 2% culture medium, 1% agarose and 10 mM phosphate buffer (pH 6.5) was solidified by dispensing on the under-lay gel and 18 hours at the optimum culture temperature of the indicator strains. Incubated for Thereafter, the size of the transparent ring formed around each sample was measured to confirm the antimicrobial activity.

As a result, RB-LBP-II-5N and RB-LBP-II-5A antimicrobial peptides showed antimicrobial activity against various indicator strains, but RB-LBP-II-5N antimicrobial peptides were found in It showed very weak antimicrobial activity against Enterobacter cloacae, Enterococcus faecalis, Edwellasiella tarda, Providencia stuartii , and showed little activity in Streptococcus mutans, Lactococcus garvieae, Streptococcus parauberis strains (Fig. 2). However, RB-LBP-II-5A, which substituted some amino acids of RB-LBP-II-5N to increase its antimicrobial activity, was used for 32 strains except Proteus mirabilis KCTC 2510, Streptococcus vestibularis KCTC 3650, and Staphylococcus haemolyticus KCTC 3341. Antimicrobial activity was observed at various concentration ranges (FIGS. 3A and 3B). Some antimicrobial peptides have only been reported to be active against Gram-positive or negative bacteria, or no fungi to fungi, but RB-LBP-II-5A antimicrobial peptides have antimicrobial activity against both Gram-positive and negative strains as well as fungi. In addition, some antibiotic resistant strains showed high antimicrobial activity, and activity showed a slight difference in activity according to species. In addition, the minimum effective concentration (MIC) was analyzed for each strain of the RB-LBP-II-5A antimicrobial peptide through the above results. The list of indicator strains used for the antimicrobial activity analysis and the minimum effect concentration (μg / ml) are shown in Table 2 below.

Type of microorganism MIC (µg / ml) Type of microorganism MIC (µg / ml) leaven Candida albicans KCCM 11282 1.4 Gram-positive bacteria Bacillus cereus KCTC 1012 17.2 Bacillus subtilis KCTC 1021 3.1 Enterococcus faecalis KCTC 3206 3.5 Lactococcus garvieae KCTC 3772 7.0 Staphylocuccus aureus KCCM 11335 4.34 Staphylocuccus aureus CCARM 0203 5.7 Staphylocuccus aureus CCARM 0204 4.7 Staphylocuccus aureus CCARM 3795 12.8 Staphylocuccus pyogenes CCARM 0206 13.3 Streptococcus mutans KCTC 3065 5.1 Streptococcus parauberis KCTC 3651 2.3 Streptococcus iniae KCTC 3657 59.1 Staphylococcus haemolyticus KCTC 3341 - Streptococcus parauberis KCTC 3551 3.0 Streptococcus vestibularis KCTC 3650 - Gram-negative bacteria Enterobacter cloacae CCARM 0252 5.5 Enterobacter cloacae KCTC 2361 2.6 Escherichia coli CCARM 0238 7.7 Escherichia coli CCARM 1A814 6.0 Escherichia coli ML35p 5.9 Edwellasiella tarda EC 10.2 Klebsiella aerogenes CCARM 0249 19.6 Klebsiella oxytoca CCARM 0248 9.8 Klebsiella pneumonia KCTC 12385 2.5 Proteus mirabilis KCTC 2510 2.2 Providencia stuartii KCTC 2569 6.3 Pseudomonas aeruginosa CCARM 0225 11.2 Pseudomonas aeruginosa KCTC 1636 3.3 Salmonella typhimurium CCARM 0240 14.0 Vibrio alginolyticus KCTC 2472 3.0 Vibrio anguillarum EC 13.0 Vibrio harveyi KCCM 40866 17.2 Vibrio parahaemolyticus KCCM 41664 12.9 Vibrio vulnificus KCCM 41665 15.0

Example 4: Evaluation of Stability against Heat and Salts

The stability against heat and salt was evaluated using the RB-LBP-II-5A antimicrobial peptide prepared according to one embodiment of the present invention.

Some antimicrobial peptides disappear after heating at 100 ° C., or antimicrobial peptides isolated from terrestrial organisms have been reported to be inhibited and disappeared by NaCl in the marine environment. RB-LBP-II-5A antimicrobial peptide was heated at 100 ° C. for 10 min and antibacterial activity at 0.5-2% NaCl condition to evaluate thermal stability and NaCl salt stability of 5A antimicrobial peptide.

As a result, RB-LBP-II-5A antimicrobial peptide showed stable antimicrobial activity without being affected by heat and NaCl (Fig. 4).

Example 5: Evaluation of Sukuchika Parasite Killing Activity

Antiparasitic activity was analyzed to evaluate the multifunctionality of the antimicrobial peptide RB-LBP-II-5A of the present invention.

Specifically, the scutica was diluted in medium to 1 × 10 ⁶ cells / 100 μl, and then 100 μl was dispensed (about 50% per well) into a 96 well plate and the sample was added to the medium. After mixing and making 100 μl concentrations, the cells were dispensed into wells containing 100 μl of parasites (sample 100 μl + parasite 100 μl = 200 μl total). Thereafter, the cells were incubated for 24 hours at 20 ° C., and morphological changes and motility changes of the Scotch caterpillar were photographed under a microscope. After the reaction was completed, 20 μl of Premix WST-1 was added per well, and the optimal reaction time after repeated measurements of absorbance (450 nm) using an ELISA reader at various reaction times (0.5, 1, 2, 3, 4 and 5 hours) Was determined. Based on the survival rate of 100% Succichica insects when treated with 0.01% acetic acid, the viability of Schizocarcci according to the antimicrobial peptide concentration of 25 ~ 200 ㎍ / ml was measured. The above formulas for experimental conditions and survival rate are as follows:

* Blank = Medium (L-15) + 0.01% acetic acid

Control group = scotchka + 0.01% acetic acid

Sample treatment group = Squatchka + peptide (1.5, 3, 6.25, 12.5, 25, 50ug / ml)

(sample-blank / control-blank) x 100 = survival rate.

As a result, the RB-LBP-II-5A antibacterial peptide was confirmed that the antiparasitic activity against the Scotchkaworm increased from 125 μg / ml to 7.5 μg / ml in addition to the antimicrobial activity. At a concentration of 15 μg / ml or higher, high killing activity was exhibited to about 85% or more of the Schizocaries but at a concentration of 7.5 μg / ml, a killing rate of about 30% was recorded (FIG. 5).

Example 6: Anticancer Activity Assay

In order to observe the anticancer effect of the multifunctional effect of the RB-LBP-II-5A antimicrobial peptide of the present invention to the uterine cancer cells (HeLa), lung cancer cells (A549) and colon cancer cells (HCT116) in the same manner as in Example 5 RB-LBP-II-5A antimicrobial peptides were treated by concentration (20-50 μg / ml) and analyzed for survival.

As a result, colorectal cancer cells (HCT116) showed a low anti-cancer activity, but in the uterine cancer cells (HeLa) and lung cancer cells (A549) very good anticancer activity was observed (Fig. 6).

In conclusion, the antimicrobial peptide of the present invention based on the post-translational protein sequence using the stone dome genome analysis results showed high antimicrobial activity against various marker strains and excellent salt and thermal stability, thus controlling various pathogenic strains and harmful strains. It can be used as a substitute for antibiotics.

Although the present invention has been described with reference to the above-described embodiments, these are merely exemplary, and it will be understood by those skilled in the art that various modifications and equivalent other embodiments are possible. Therefore, the true technical protection scope of the present invention will be defined by the technical spirit of the appended claims.

<110> Republic of Korea represented by National Fisheries Research & Development Institute <120> Antimicrobial peptide derived from rock bream and uses <130> PD17-5581 <160> 3 <170> KopatentIn 2.0 <210> 1 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> RB-LBP-II-5 <400> 1 Pro Gln Ile Ala Gln Arg Phe Pro Gly Leu Met Met Lys Leu Leu Val   1 5 10 15 <210> 2 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> RB-LBP-II-5N <400> 2 Arg Phe Pro Gly Leu Met Met Lys Leu Leu   1 5 10 <210> 3 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> RB-LBP-II-5A <400> 3 Arg Phe Leu Lys Leu Met Met Lys Leu Leu   1 5 10

Claims (11)

An antimicrobial peptide, consisting of the amino acid sequence set forth in SEQ ID NO: 3 (RFLKLMMKLL). The method of claim 1,
Antimicrobial peptides amidated C-terminus. An antimicrobial composition comprising the antimicrobial peptide of claim 1 as an active ingredient. The method of claim 3,
Antimicrobial composition having antimicrobial activity against gram negative bacteria, gram positive bacteria or fungi. The method of claim 4, wherein
The Gram-positive bacteria are B. cereus, B. subtilis, S. mutans, S. iniae, S. aureus, L. garvieae, S. parauberis, S. aureus 0203 , S. aureus 0204 , S. pyogenes 0206, or S. aureus . The antimicrobial composition of aureus 3795. The method of claim 4, wherein
The Gram-negative bacteria are E. coli, P. aeruginosa, V. alginoalyticus, V. anguillarum, V. parahaemolyticus, V. vulnificus, V. harveyi, K. pneumoniae, E. tzarda, P. stuartii, E. coli ML35p, P antimicrobial composition which is aeruginosa 0225, E. coli 0238, S. typhimurium 0240, K. oxytoca 0248, K. aerogenes 0249, E. cloacae 0252, or E. coli 1A814. The method of claim 4, wherein
The fungus is C. albicans , antimicrobial composition. Feed additive containing the antimicrobial peptide of Claim 1 or 2 as an active ingredient. The antimicrobial peptide of Claim 1 or Claim 2 containing as an active ingredient, the composition for controlling Scotch caterpillars. An anticancer composition comprising the antimicrobial peptide of claim 1 as an active ingredient. The method of claim 10,
The cancer is uterine cancer or lung cancer, anticancer composition.

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