Bacillus cereus ATP synthase beta subunit antibacterial peptide BaD9 and application thereof

Abstract

The application discloses a bacillus cereus ATP synthase beta subunit antibacterial peptide BaD9, and the amino acid sequence of the bacillus cereus ATP synthase beta subunit antibacterial peptide BaD9 is KLVVHRARRIQFFLSQNFH, and the molecular weight of the antibacterial peptide is 2396.831 Dalton. The antibacterial peptide has obvious inhibiting effect on Vibrio parahaemolyticus, Staphylococcus aureus and Staphylococcus haemolyticus, and can be used for preparing medicines for preventing or inhibiting Vibrio parahaemolyticus, Staphylococcus aureus and Staphylococcus haemolyticus infection, food preservatives or feed additives.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to an antimicrobial peptide BaD9, which is a β subunit of Bacillus cereus ATP synthase, and its applications. Background Technology

[0002] Vibrio parahaemolyticus ( Vibrio parahaemolyticus Vibrio parahaemolyticus (Staphylococcus aureus) belongs to the genus Vibrio, is a Gram-negative bacterium, and is a major pathogen in aquaculture. It is also one of the five most common foodborne pathogens. Consuming seafood contaminated with pathogenic Vibrio parahaemolyticus can cause acute gastroenteritis. Symptoms include dizziness, diarrhea, abdominal cramps, vomiting, and fever; in severe cases, it can lead to dehydration and coma. Staphylococcus aureus Staphylococcus aureus (S. aureus) is a Gram-positive bacterium belonging to the genus Staphylococcus. It is a common food contaminant and one of the five major foodborne pathogens. It can produce enterotoxins, causing food poisoning. Patients infected with Staphylococcus aureus may experience nausea, vomiting, and spasmodic pain in the upper middle abdomen, followed by diarrhea, with vomiting being the most prominent symptom. Hemolytic Staphylococcus aureus (S. aureus) Staphylococcus haemolyticus It belongs to the genus Staphylococcus, is a Gram-positive bacterium, and is a normal colonizing bacterium on the human body surface. Its carrier rate is second only to Staphylococcus epidermidis. It is a typical opportunistic pathogen that often causes infection in patients with weakened immune systems and patients using indwelling or implanted foreign bodies. It is an important pathogen causing hospital-acquired infections.

[0003] The use of antibiotics to treat bacterial diseases is a major advancement in human medical history. However, due to antibiotic overuse, bacterial resistance to antibiotics is becoming increasingly serious, posing a significant threat to human health. Unlike antibiotics, which interfere with the metabolic processes of pathogenic microorganisms, antimicrobial peptides mostly exert their antibacterial effects by disrupting the cell membranes of microorganisms and inducing leakage of cell contents, thus being least susceptible to bacterial resistance. Therefore, antimicrobial peptides are considered a good alternative to antibiotics.

[0004] Antimicrobial peptides (AMPs) are a class of small molecule peptides widely distributed in organisms, forming part of the body's first line of defense against pathogens and effectively resisting the invasion of pathogenic microorganisms. Antimicrobial peptides possess broad-spectrum anti-biofilm activity, are beneficial in regulating host immune responses, and are less likely to induce drug resistance, thus exhibiting unique advantages in the treatment of infectious diseases and showing promise as ideal anti-infective drugs. Currently, antimicrobial peptides have been discovered and isolated from bacteria, fungi, higher plants, and animals.

[0005] Bacillus cereus is a Gram-positive, rod-shaped, spore-forming facultative anaerobic bacterium frequently found in soil, water, and animal intestines. It can produce toxins that cause various types of gastrointestinal or other diseases. Bacillus can also antagonize fungal pathogens through antibacterial, nutrient competition, site repulsion, parasitism, or induction. It can also produce various active substances, such as low-molecular-weight peptides, lipopeptide antibiotics, and antimicrobial proteins, which can effectively inhibit the growth of various pathogenic microorganisms, possessing significant research and application value. The ATP synthase subunit beta (AtpD, ATP synthase β subunit) gene encodes a subunit of mitochondrial ATP synthase. Mitochondrial ATP synthase catalyzes ATP synthesis, utilizing the electrochemical gradient of protons crossing the inner membrane during oxidative phosphorylation. Studies have shown that it has an effective protective effect against psychrophilic bacteria. Therefore, identifying antimicrobial peptides with antimicrobial activity from the Bacillus cereus ATP synthase β subunit and exploring their antibacterial mechanism is a pressing technical challenge in this field. Summary of the Invention

[0006] The purpose of this invention is to overcome the shortcomings of the prior art and provide a Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9 and its application, thus solving the problems in the background art mentioned above.

[0007] One of the technical solutions adopted by the present invention to solve its technical problem is: providing a Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9, the amino acid sequence of which is shown in SEQ ID NO: 1: KLVVHRARRIQFFLSQNFH The antimicrobial peptide BaD9 has a molecular weight of 2396.831 Daltons, a positive charge of +4.5, and a total hydrophobicity of 47%. APD3 analysis suggests that the peptide may form an α-helix.

[0008] The second technical solution adopted by the present invention to solve its technical problem is: providing the application of Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9 in the preparation of antimicrobial drugs, which are used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus.

[0009] The third technical solution adopted by the present invention to solve its technical problem is: to provide an antibacterial drug, the effective ingredient of which includes the antimicrobial peptide BaD9 of Bacillus cereus ATP synthase β subunit, wherein the amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0010] In a preferred embodiment of the present invention, the active ingredient of the antibacterial drug is the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase, and the amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0011] In a preferred embodiment of the present invention, the antibacterial drug is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

[0012] The fourth technical solution adopted by the present invention to solve its technical problem is: providing the application of Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9 in the preparation of food preservatives, which are used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus.

[0013] The fifth technical solution adopted by the present invention to solve its technical problem is: providing a food preservative, the effective component of which includes Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9, the amino acid sequence of said antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0014] In a preferred embodiment of the present invention, the active ingredient of the food preservative is Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9, and the amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0015] In a preferred embodiment of the present invention, the food preservative is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

[0016] The sixth technical solution adopted by the present invention to solve its technical problem is: providing the application of Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9 in the preparation of aquatic feed additives, which are used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus.

[0017] The seventh technical solution adopted by the present invention to solve its technical problem is: to provide an aquatic feed additive, the effective ingredient of which includes Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9, the amino acid sequence of said antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0018] In a preferred embodiment of the present invention, the effective component of the aquatic feed additive is Bacillus cereus ATP synthase β subunit antimicrobial peptide BaD9, and the amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO: 1.

[0019] In a preferred embodiment of the present invention, the aquatic feed additive is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

[0020] This invention utilizes software and websites such as APD3, Pymol2.0, and SWISS-MODEL to perform bioinformatics prediction on the protein sequence of the ATP synthase β subunit protein of Bacillus cereus, which has antibacterial activity, to obtain a new amino acid sequence polypeptide, BaD9. The antibacterial activity of BaD9 against Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus was studied. Taking Vibrio parahaemolyticus as an example, the degree of damage to the bacterial cell membrane by BaD9 was observed using flow cytometry and PI staining. Simultaneously, genomic DNA of Vibrio parahaemolyticus was extracted to verify its effect on bacterial DNA. Experimental results show that this peptide has a significant inhibitory effect on Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus. The antimicrobial peptide BaD9 of this invention is a cationic antimicrobial peptide. Its positive charge can bind to the anionic bacterial membrane through electrostatic interaction, thereby adsorbing onto the bacterial membrane, changing the cell membrane potential, causing cell membrane damage, and exerting its antibacterial effect. Furthermore, the α-helical structure of the antimicrobial peptide BaD9 can induce the formation of transmembrane pores on the bacterial surface, thereby facilitating the peptide's passage through the cell membrane or causing leakage of liposomes and biomolecules within the cell. Therefore, regardless of whether the bacteria are Gram-positive or Gram-negative, the antimicrobial peptide BaD9 binds to its cell membrane surface in the same way. Combined with MBC and TIME-KILL time results, this antimicrobial peptide exhibits inhibitory effects against Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

[0021] The antimicrobial peptide BaD9 will damage bacteria through the following mechanisms: BaD9 can penetrate the cell membrane, bind to bacterial genomic DNA, inhibit bacterial DNA synthesis, and thus lead to bacterial death.

[0022] The antimicrobial peptides of the present invention can be synthesized using methods known to those skilled in the art, such as solid-phase synthesis, and purified using methods known to those skilled in the art, such as high-performance liquid chromatography.

[0023] Implementing this invention has the following beneficial effects: This invention, through screening, discovered a novel polypeptide, BaD9, with a novel amino acid sequence. Studies have shown that BaD9 exhibits significant inhibitory effects against Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci. Its antibacterial mechanism involves first penetrating the bacterial cell membrane, then binding to the bacterial genomic DNA, inhibiting bacterial DNA synthesis, thereby leading to bacterial death. The antimicrobial peptide BaD9 of this invention can be formulated into antimicrobial drugs or aquatic feed additives for the prevention or treatment of diseases caused by Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci. Attached Figure Description

[0024] Figure 1 This is a schematic diagram of the structure of the antimicrobial peptide BaD9.

[0025] Figure 2 This is a control diagram showing the determination of the minimum bactericidal concentration (MBC) of the antimicrobial peptide BaD9 against Vibrio parahaemolyticus.

[0026] The concentrations of the antimicrobial peptide BaD9 in Figure AI were 0 μg / mL, 500 μg / mL, 250 μg / mL, 125 μg / mL, 62.5 μg / mL, 31.25 μg / mL, 15.625 μg / mL, 7.813 μg / mL, and 3.9 μg / mL, respectively. Figure 3 This is a control chart showing the determination of the minimum bactericidal concentration (MBC) of the antimicrobial peptide BaD9 against hemolytic staphylococci.

[0027] The concentrations of the antimicrobial peptide BaD9 in Figure AI were 0 μg / mL, 500 μg / mL, 250 μg / mL, 125 μg / mL, 62.5 μg / mL, 31.25 μg / mL, 15.625 μg / mL, 7.813 μg / mL, and 3.9 μg / mL, respectively. Figure 4 This is a control chart showing the determination of the minimum bactericidal concentration (MBC) of the antimicrobial peptide BaD9 against Staphylococcus aureus.

[0028] The concentrations of the antimicrobial peptide BaD9 in Figure AI were 0 μg / mL, 500 μg / mL, 250 μg / mL, 125 μg / mL, 62.5 μg / mL, 31.25 μg / mL, 15.625 μg / mL, 7.813 μg / mL, and 3.9 μg / mL, respectively. Figure 5 The time-dependent killing kinetics curve of the antimicrobial peptide BaD9 against Vibrio parahaemolyticus is shown.

[0029] Figure 6 The time-dependent killing kinetics curve of the antimicrobial peptide BaD9 against hemolytic staphylococci.

[0030] Figure 7 The time-dependent killing kinetics curve of the antimicrobial peptide BaD9 against Staphylococcus aureus.

[0031] Figure 8 The effect of the antimicrobial peptide BaD9 on the cell membrane permeability (protein leakage) of Vibrio parahaemolyticus.

[0032] Figure 9 The effect of the antimicrobial peptide BaD9 on the cell membrane permeability of Vibrio parahaemolyticus.

[0033] Figure 10 The image shows the circular dichroism chromatograms of the antimicrobial peptide BaD9 under different conditions.

[0034] Figure 11 This is a gel electrophoresis image of the antimicrobial peptide BaD9 and Vibrio parahaemolyticus genomic DNA.

[0035] Among them, the mass ratios of antimicrobial peptide BaD9 / DNA for bands 1-9 are 25 / 4, 25 / 8, 25 / 10, 25 / 12, 25 / 14, 25 / 16, 25 / 18, 25 / 20, and 25 / 25, respectively. Band 10 represents the control group DNA.

[0036] Figure 12 This is a circular dichroism chromatogram showing the changes in the antimicrobial peptide BaD9 after it binds to the genomic DNA of Vibrio parahaemolyticus. Detailed Implementation

[0037] To better understand the present invention, the present invention will be further described in detail below with reference to the embodiments and accompanying drawings. However, those skilled in the art will understand that the following embodiments are not intended to limit the scope of protection of the present invention, and any changes and variations made on the basis of the present invention are within the scope of protection of the present invention.

[0038] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0039] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0040] Example 1: Screening of antimicrobial peptide BaD9 Bioinformatics prediction was performed on the protein sequence of the ATP synthase β subunit protein with antibacterial activity in Bacillus cereus using software and websites such as APD3, Pymol2.0, and Swiss-Model. The amino acid sequence was analyzed for charge and hydrophobicity using APD3 (Table 1), and the three-dimensional structure was predicted using Pymol2.0 and Swiss-Model (e.g., ...). Figure 1 As shown in the figure, the amino acid sequence KLVVHRARRIQFFLSQNFH with the best antibacterial properties was finally screened out and chemically synthesized (by Beijing Zhongke Yaguang Biotechnology Co., Ltd.), and its antibacterial activity was verified.

[0041] Table 1: Predictive analysis of antimicrobial peptides in Bacillus cereus peptide sequence Molecular weight (Da) hydrophobicity Net charge Source (protein) KLVVHRARRIQFFLSQNFH 2396.831 47% +4.5 ATP synthase subunit beta Example 2 Determination of the minimum bactericidal concentration (MBC) of the antimicrobial peptide BaD9 Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic Staphylococcus (Vibrio parahaemolyticus ATCC17802, Staphylococcus aureus ATCC27217, and hemolytic Staphylococcus JCM2416, all strains preserved in our laboratory) were cultured at 37°C for 12 hours to the logarithmic growth phase, and then diluted to 10 μL in 0.01 M pH 7.2 phosphate buffer. 5-6 CFU / mL. The antimicrobial peptide BaD9 was dissolved in phosphate buffer, and an equal volume was mixed with bacteria at 37°C and incubated for 2 h. The minimum inhibitory concentration (MBC) is the lowest concentration of antimicrobial peptide that can kill bacteria after incubation at 37°C. For example... Figures 2-4 As shown, the minimum bactericidal concentrations (MBCs) of the antimicrobial peptide BaD9 against Vibrio parahaemolyticus, Staphylococcus aureus, and Staphylococcus aureus were 3.90625 µg / mL, 7.8125 µg / mL, and 7.8125 µg / mL, respectively.

[0042] Example 3: Determination of the time-kill curve of antimicrobial peptide BaD9 Vibrio parahaemolyticus, Staphylococcus hemolyticus, and Staphylococcus aureus were cultured at 37°C for 12 h until they reached the logarithmic growth phase, and then diluted to a final concentration in 0.01M pH 7.2 phosphate buffer. 5-6 CFU / mL. A 1×MBC concentration peptide was mixed with an equal volume of bacteria at 37℃ and incubated separately. Samples were taken every 30 minutes and plated. After overnight incubation at 37℃, the total colony count was recorded. The results showed that the antimicrobial peptides could kill Vibrio parahaemolyticus, hemolytic Staphylococcus, and Staphylococcus aureus within 0.5 hours. (e.g., Figures 5-7 (As shown) Example 4: Effect of antimicrobial peptide BaD9 on cell membrane permeability of Vibrio parahaemolyticus Protein leakage: 200 μL of Vibrio parahaemolyticus strain stored at -20℃ was inoculated into LB medium and cultured at 37℃ and 200 r / h for 10 h until the bacteria reached the logarithmic growth phase. After preparing a bacterial suspension in 0.01 M phosphate buffer, it was mixed with antimicrobial peptide BaD9 at concentrations of 1×MBC, 2×MBC, and 4×MBC in equal proportions. The OD280 nm concentration was measured every 10 min between 0 and 2 h using a multi-functional microplate reader to assess the amount of protein leakage. An equal volume of 0.01 M phosphate buffer mixed with bacteria in equal proportions served as a blank control (e.g., ...). Figure 8 (As shown).

[0043] Detection of absorbance at 280 nm can be used to estimate the amount of protein leaking from the cytoplasm. For example... Figure 8 As shown, with the increase of antimicrobial peptide BaD9 concentration, the protein content in the culture medium treated with antimicrobial peptide BaD9 increased in a dose-dependent manner.

[0044] Flow cytometry: Vibrio parahaemolyticus was cultured at 37°C for 12 h to the logarithmic growth phase. 1 mL of bacterial suspension was centrifuged at 12000 r / min for 1 min in a 1.5 mL centrifuge tube, the supernatant was removed, and the suspension was resuspended in 1 mL of sterile 0.01 M phosphate buffer. This process was repeated three times. After preparing the bacterial suspension in 0.01 M phosphate buffer, the antimicrobial peptide BaD9 at concentrations of 1×MBC and 2×MBC was mixed with the bacterial suspension in equal proportions (greater than 200 μL) and incubated at 37°C for 2 h. 0.01 M phosphate buffer was used as a blank control. Pyridine iodide (PI) was used as the fluorescent dye. After incubation, an equal volume of 50 μg / mL PI dye was added to the mixture, and the mixture was incubated at 4°C for 15 min before being loaded onto a flow cytometer for detection. Each tube was designed to collect at least 10 samples. 4 One bacterial cell. A data collection and analysis protocol was established, and the sample was analyzed by detecting the scattered light signal (SC) and the propidium iodide fluorescence signal (PI) (e.g., Figure 9 (As shown).

[0045] like Figure 9 As shown, the mortality rate of Vibrio parahaemolyticus in the control group was only 9.95%, indicating that the cell membrane structure was intact, preventing PI from crossing the cell membrane and entering the cell, resulting in a low bacterial mortality rate. The mortality rate of Vibrio parahaemolyticus significantly increased to 89.8% with the addition of 1×MBC concentration of antimicrobial peptide BaD9. In the experimental group with 2×MBC concentration of antimicrobial peptide, the mortality rate of Vibrio parahaemolyticus was slightly higher than that with 1×MBC, reaching 94.6%. This demonstrates a positive correlation between the concentration of antimicrobial peptide BaD9 and the mortality rate of Vibrio parahaemolyticus, exhibiting a dose-dependent increase.

[0046] Example 5: Determination of the secondary structure of the antimicrobial peptide BaD9 by circular dichroism chromatography The average residue molar ellipticity of the antimicrobial peptide BaD9 was determined using a Jasco 810 spectropolarimeter (Jasco, Tokyo) CD at 25 °C and a scan rate of 100 nm / min. The antimicrobial peptide BaD9 was dissolved in 0.01 M PBS and 25 mM sodium dodecyl sulfate (SDS) to a final concentration of 0.20 mg / mL, and its spectrum was scanned twice from 180 to 280 nm.

[0047] like Figure 10As shown, in PBS, the antimicrobial peptide BaD9 exhibits a positive peak at 198 nm and a negative peak at 225 nm, indicating that its secondary conformation in PBS is primarily β-sheet. In SDS, a positive peak is observed at 190 nm, while two negative peaks appear at 211 nm and 220 nm, suggesting that its secondary conformation in SDS is primarily α-helix. This indicates that the antimicrobial peptide may undergo structural transformation upon contact with the bacterial cell membrane.

[0048] Example 6: Interaction between antimicrobial peptide BaD9 and bacterial DNA The interaction between the antimicrobial peptide BaD9 and the genomic DNA of Vibrio parahaemolyticus was investigated using the DNA gel retardation assay. Vibrio parahaemolyticus was cultured in 50 mL of nutrient broth at 37°C for 12 h. The interaction was studied using the bacterial optical density ratio (OD) at 260 and 280 nm. 260 / OD 280 The purity of the extracted genomic DNA was evaluated using a ≥ 1.90 ppm. Next, 10 µL of DNA (20 ng / µL) was mixed with the antimicrobial peptide BaD9 at 25°C to achieve BaD9 / DNA mass ratios of 25 / 4, 25 / 8, 25 / 10, 25 / 12, 25 / 14, 25 / 16, 25 / 18, 25 / 20, 25 / 25, and 0, respectively. The mixture was then incubated at 37°C for 1.5 h. Eight µL of each mixture was then electrophoresed on a 1% agarose gel, and gel retardation was observed under UV irradiation using a GelDoc XR gel imaging system (Bio-Rad, USA).

[0049] like Figure 11 As shown, when the ratios were 25 / 4, 25 / 8, 25 / 10, and 25 / 12, the DNA bands did not escape but remained clearly in the sample wells. When the ratios were 25 / 14, 25 / 16, 25 / 18, 25 / 20, and 25 / 25, the clarity and brightness of the bands increased significantly as the ratio decreased. The gel electrophoresis image of the control group, Vibrio parahaemolyticus genomic DNA, showed the brightest and clearest band. This indicates that when the antimicrobial peptide BaD9 / DNA mass ratio was 25 / 4, 25 / 8, 25 / 10, and 25 / 12, the DNA was blocked and destroyed, and the bands did not escape. As the concentration of the antimicrobial peptide decreased, the bands gradually became clearer.

[0050] Example 7: Effect of the interaction between the antimicrobial peptide BaD9 and bacterial DNA on its secondary structure The average residue molar ellipticity of the antimicrobial peptide BaD9 was determined again using a Jasco 810 spectropolarimeter (Jasco, Tokyo) CD at 25°C and a scan rate of 100 nm / min. The antimicrobial peptide BaD9 was dissolved in 25 mM sodium dodecyl sulfate (SDS) to a final concentration of 0.20 mg / mL. Vibrio parahaemolyticus DNA (20 ng / μL) was mixed with the antimicrobial peptide BaD9 and incubated at 25°C for 2 h. The spectrum was then scanned twice from 180 to 280 nm. Figure 12 (As shown).

[0051] After incubation with the DNA, the secondary structure of the antimicrobial peptide BaD9 changed. The positive peak changed from 190 nm to 195 nm, and the negative peak changed from a double negative peak at 211 nm and 220 nm to a single negative peak at 217 nm. The secondary structure changed from an α-helix structure to a β-sheet structure, and the spectrum of the antimicrobial peptide BaD9 was enlarged overall. This indicates that the antimicrobial peptide BaD9 interacts with the genomic DNA of Vibrio parahaemolyticus and can cause changes in the secondary structure conformation of the antimicrobial peptide.

[0052] In summary, this invention provides a novel antimicrobial peptide, BaD9. The minimum bactericidal concentrations (MBCs) of BaD9 against Vibrio parahaemolyticus, Staphylococcus aureus, and Staphylococcus aureus are 3.90625 µg / mL, 7.8125 µg / mL, and 7.8125 µg / mL, respectively. All three bacteria are killed within 0.5 hours, demonstrating a strong inhibitory effect on BaD9. Studies have also found that BaD9 can penetrate bacterial cell membranes, bind to bacterial genomic DNA, inhibit bacterial DNA synthesis, and thus lead to bacterial death.

[0053] While specific embodiments of the present invention have been described above, those skilled in the art should understand that the specific embodiments described are merely illustrative and not intended to limit the scope of the present invention. Equivalent modifications and variations made by those skilled in the art in accordance with the spirit of the present invention should be covered within the scope of protection of the claims of the present invention.

Claims

1. An antimicrobial peptide BaD9, a β subunit of Bacillus cereus ATP synthase, the amino acid sequence of which is shown in SEQ ID NO:

1.

2. The application of the antimicrobial peptide BaD9, a β-subunit of Bacillus cereus ATP synthase as described in claim 1, in the preparation of antimicrobial drugs, characterized in that: The antibacterial drug is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

3. An antibacterial drug, characterized in that: Its active ingredient includes the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO:

1.

4. The antibacterial drug as described in claim 3, characterized in that: Its active ingredient is the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO:

1.

5. The application of the antimicrobial peptide BaD9, a β-subunit of Bacillus cereus ATP synthase as described in claim 1, in the preparation of a food preservative, characterized in that: The food preservative is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

6. A food preservative, characterized in that: Its active ingredient includes the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO:

1.

7. The food preservative as described in claim 6, characterized in that: Its active ingredient is the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO:

1.

8. The application of the antimicrobial peptide BaD9, a β-subunit of Bacillus cereus ATP synthase as described in claim 1, in the preparation of aquatic feed additives, characterized in that: The aquatic feed additive is used to inhibit and / or kill one or more of Vibrio parahaemolyticus, Staphylococcus aureus, and hemolytic staphylococci.

9. An aquatic feed additive, characterized in that: Its active ingredient includes the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO:

1.

10. The aquatic feed additive as described in claim 9, characterized in that: Its active ingredient is the antimicrobial peptide BaD9, which is the β subunit of Bacillus cereus ATP synthase. The amino acid sequence of the antimicrobial peptide BaD9 is SEQ ID NO: 1.

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