An antimicrobial peptide HSP-M4 constructed based on rational molecular design and its preparation method and application

Abstract

The present application relates to the field of biomedicine technology, and in particular to an antimicrobial peptide HSP-M4 constructed based on rational molecular design, a preparation method and an application thereof; the amino acid sequence of the antimicrobial peptide HSP-M4 is shown in SEQ ID NO.1, and the antimicrobial peptide HSP-M4 is a derivative peptide of the antimicrobial peptide HSP-1; a low-toxic antimicrobial peptide HSP-M4 is formed by molecular modification of the natural antimicrobial peptide HSP-1, optimization and modification of the amino acid sequence of the antimicrobial peptide HSP-1, and secondary structure prediction, antimicrobial activity detection and cytotoxicity detection, and the antimicrobial peptide HSP-M4 has significantly improved bactericidal activity against Escherichia coli, Staphylococcus aureus and Candida albicans compared with the natural antimicrobial peptide HSP-1.

Description

Technical Field

[0001] The present application relates to the field of biomedicine technology, and in particular to an antimicrobial peptide HSP-M4 constructed based on rational molecular design, and a preparation method and application thereof. Background Art

[0002] Antibiotics, as the greatest invention of the 20th century, have made an important contribution to human resistance to pathogen infection. However, with the abuse of antibiotics, clinical findings have shown that multidrug resistance in cells is prominent, which will lead to super bacteria. The current pace of antibiotic research and development is far behind the evolution of bacterial resistance. This is because the antibiotic bactericidal mechanism mainly has a specific receptor, such as DNA, pumps on the cell membrane, peptidoglycan layer, etc., while antimicrobial peptides do not have a specific target receptor and mainly destroy the cell membrane structure through physical action. The unique bactericidal mechanism of antimicrobial peptides is that the positively charged and amphiphilic antimicrobial peptides electrostatically adsorb the negative charges in the bacterial cell membrane, and the subsequent hydrophobic sequence enters the phospholipid bilayer of the cell membrane, destroying the cell membrane structure and interfering with the osmotic pressure of the cell, which ultimately leads to bacterial rupture and death. Therefore, antimicrobial peptides are not easy to develop resistance. Based on this, antimicrobial peptides are expected to become one of the most effective directions for solving the problem of antibiotic resistance.

[0003] Antimicrobial peptides are a type of defensive peptide produced in organisms against infection by external pathogens, and exist in a variety of organisms. Therefore, according to the source of antimicrobial peptides, they can be divided into bacterial antimicrobial peptides, animal antimicrobial peptides, plant antimicrobial peptides, etc. The amino acid sequence length of general antimicrobial peptides is about 10-50 amino acids, and the sequence often contains positively charged amino acid residues (for example, lysine, arginine) and hydrophobic amino acids (such as aromatic amino acids). This characteristic is the structural basis for the binding of antimicrobial peptide molecules to target cells. Among them, the secondary structure of antimicrobial peptides is mostly α-helix, and this structure is the main structure for antimicrobial peptides to maintain activity. At present, the biological activity of antimicrobial peptides is relatively broad, which can inhibit microorganisms such as G+, G-, fungi and viruses, and can also inhibit the growth of parasites and tumor cells. In addition, it also plays an important role in food preservation.

[0004] HSP-1 (Hylaseptin-1) is a natural antimicrobial peptide originally isolated from Rana sylvatica (commonly found in wetland areas in northern and northeastern Brazil). Activity experiments have shown that HSP-1 does not show obvious hemolytic activity and has less toxic side effects, but has a mild inhibitory effect on Candida albicans, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa, and has a better inhibitory effect on fungi. Based on this, HSP-1 is often selected as an antimicrobial peptide modification object. However, since the inhibitory effect of antimicrobial peptides on bacteria such as Candida albicans, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa is not obvious, how to develop an HSP-1 antimicrobial peptide to improve the inhibitory effect of antimicrobial peptides on bacteria such as Candida albicans, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa is a technical problem that needs to be solved urgently. Summary of the invention

[0005] The present application provides an antimicrobial peptide HSP-M4 constructed based on rational molecular design, as well as a preparation method and application, to solve the technical problem in the prior art that the inhibitory effect of HSP-1 antimicrobial peptide on Candida albicans, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa is not obvious.

[0006] In a first aspect, the present application provides an antimicrobial peptide HSP-M4 constructed based on rational molecular design, the amino acid sequence of the antimicrobial peptide HSP-M4 is shown in SEQ ID NO.1, and the antimicrobial peptide HSP-M4 is a derivative peptide of the antimicrobial peptide HSP-1.

[0007] Optionally, the C-terminus of the antimicrobial peptide HSP-M4 is an acetamidate terminus.

[0008] In a second aspect, the present application provides a method for preparing the antimicrobial peptide HSP-M4 described in the first aspect, the method comprising:

[0009] Optimize the amino acid sequence of wild-type HSP-1 to obtain derivative peptides of multiple antimicrobial peptides;

[0010] The secondary structure of the derived peptide is predicted, and then the antibacterial activity test and cytotoxicity test are performed to determine the best derived peptide.

[0011] Optionally, the optimization includes optimizing with the goal of optimizing the net charge number and hydrophobicity.

[0012] Optionally, the optimization further comprises modifying the amino acid sequence of the wild-type HSP-1, wherein the modification comprises modifying the wild-type HSP-1 by truncating the N-terminus of the amino acid sequence of the wild-type HSP-1 and / or truncating the C-terminus of the amino acid sequence of the wild-type HSP-1 and / or extending the α-helix of the amino acid sequence of the wild-type HSP-1.

[0013] In a third aspect, the present application provides a use of the antimicrobial peptide HSP-M4 described in the first aspect in the preparation of a pharmaceutical composition, wherein the pharmaceutical composition is used for antibacterial or antifungal purposes.

[0014] Optionally, the bacteria include Escherichia coli and / or Staphylococcus aureus; and / or,

[0015] The fungi include Candida albicans.

[0016] In a fourth aspect, the present application provides an antibacterial drug, which comprises the active ingredient of the antimicrobial peptide HSP-M4 described in the first aspect.

[0017] Optionally, the antibacterial drug is a drug for treating bacterial infection and / or fungal infection.

[0018] Optionally, the bacterial infection is caused by Escherichia coli and / or Staphylococcus aureus; and / or,

[0019] The fungal infection is caused by Candida albicans.

[0020] The above technical solution provided by the embodiment of the present application has the following advantages compared with the prior art:

[0021] The embodiment of the present application provides an antimicrobial peptide HSP-M4 constructed based on rational molecular design, which forms a low-toxic antimicrobial peptide HSP-M4 by molecular modification of the natural antimicrobial peptide HSP-1, optimization and modification of the amino acid sequence of the antimicrobial peptide HSP-1, and secondary structure prediction, antimicrobial activity detection and cytotoxicity detection. At the same time, the antimicrobial peptide HSP-M4 has significantly improved bactericidal activity against Escherichia coli, Staphylococcus aureus and Candida albicans compared with the natural antimicrobial peptide HSP-1. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the present application and, together with the description, serve to explain the principles of the present application.

[0023] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative labor.

[0024] Figure 1 A schematic diagram of the antimicrobial peptide HSP-M4 provided in the examples of the present application;

[0025] Figure 2 A schematic diagram of the preparation method provided in the embodiment of the present application;

[0026] Figure 3 The spiral projection model of the HSP-1 series antimicrobial peptides provided in the examples of the present application analyzed by heliQuest software,

[0027] Among them, the hydrophobic surface Figure 3 As shown by the middle arrow, its hydrophobic distance reaches a maximum value of 0.793;

[0028] Figure 4 The secondary structure model of the HSP-1 series antimicrobial peptides provided in the examples of the present application predicted by the I-TASSER software;

[0029] Figure 5 A schematic diagram of the results of the hemolysis rate of red blood cells of the HSP-1 series of antimicrobial peptides provided in the examples of the present application;

[0030] Figure 6 This is a schematic diagram of the HEK293 cell toxicity results of the HSP-1 series antimicrobial peptides provided in the examples of the present application. DETAILED DESCRIPTION

[0031] In order to make the purpose, technical solution and advantages of the embodiments of the present application clearer, the technical solution in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of this application.

[0032] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in this application can be purchased from the market or prepared by existing methods.

[0033] The embodiment of the present application provides an antimicrobial peptide HSP-M4 constructed based on rational molecular design. The amino acid sequence of the antimicrobial peptide HSP-M4 is shown in SEQ ID NO.1. The antimicrobial peptide HSP-M4 is a derivative peptide of the antimicrobial peptide HSP-1.

[0034] In some optional embodiments, the C-terminus of the antimicrobial peptide HSP-M4 is an acetamidate terminus.

[0035] In the examples of the present application, the C-terminus of the antimicrobial peptide HSP-M4 is limited to an acetamidate terminus, which can effectively improve the efficiency of the antimicrobial peptide HSP-M4 entering and exiting the cell membrane, thereby improving the bactericidal effect.

[0036] like Figure 1 and Figure 2As shown, based on a general inventive concept, the present application embodiment provides a method for preparing the antimicrobial peptide HSP-M4, the method comprising:

[0037] S1. Optimize the amino acid sequence of wild-type HSP-1 to obtain derivative peptides of multiple antimicrobial peptides;

[0038] S2. Predict the secondary structure of the derivative peptide, and then perform antibacterial activity test and cytotoxicity test to determine the best derivative peptide.

[0039] In the examples of the present application, by optimizing the amino acid sequence of wild-type HSP-1, not only the net charge number and hydrophobicity of the antimicrobial peptide can be optimized, but also all aspects affecting the antimicrobial activity of the antimicrobial peptide can be comprehensively considered from the perspective of the helical structure, thereby designing antimicrobial peptides with effective antimicrobial activity against Candida albicans, Staphylococcus aureus, and Escherichia coli.

[0040] The method is a method for preparing the antimicrobial peptide HSP-M4. The specific amino acid sequence of the antimicrobial peptide HSP-M4 can refer to the above embodiment. Since the method adopts part or all of the technical solutions of the above embodiment, it at least has all the beneficial effects brought by the technical solutions of the above embodiment, which will not be repeated here.

[0041] In some optional embodiments, the optimization includes optimizing with the goal of optimizing the net charge number and hydrophobicity.

[0042] In the examples of the present application, the specific optimization target is limited. Since the net charge number and hydrophobicity carried by the polypeptide have a certain influence on the activity of the antimicrobial peptide, the optimization is considered for both, which can effectively ensure that the designed antimicrobial peptide has strong antimicrobial performance.

[0043] In some optional embodiments, the optimization further comprises modifying the amino acid sequence of the wild-type HSP-1, wherein the modification comprises modifying the wild-type HSP-1 by truncating the N-terminus of the amino acid sequence of the wild-type HSP-1 and / or truncating the C-terminus of the amino acid sequence of the wild-type HSP-1 and / or extending the α-helix of the amino acid sequence of the wild-type HSP-1.

[0044] In the examples of the present application, the specific method of modification is limited. The helical structure of the antimicrobial peptide is crucial to the antimicrobial activity of the antimicrobial peptide. Therefore, by truncating the two ends of the amino acid sequence of the wild-type HSP-1 or extending the α-helix, the derivative peptide of the antimicrobial peptide HSP-1 with the strongest antimicrobial performance is screened.

[0045] Based on a general inventive concept, an embodiment of the present application provides a use of the antimicrobial peptide HSP-M4 in preparing a pharmaceutical composition, wherein the pharmaceutical composition is used for antibacterial or antifungal purposes.

[0046] This application is realized based on the above-mentioned antimicrobial peptide HSP-M4. The specific amino acid sequence of the antimicrobial peptide HSP-M4 can refer to the above-mentioned embodiment. Since this application adopts part or all of the technical solutions of the above-mentioned embodiment, it has at least all the beneficial effects brought by the technical solutions of the above-mentioned embodiment, which will not be described one by one here.

[0047] In some optional embodiments, the bacteria include Escherichia coli and / or Staphylococcus aureus; and / or,

[0048] The fungi include Candida albicans.

[0049] In the examples of the present application, specific bacteria and fungi species are limited because the antimicrobial peptide HSP-M4 has more effective antimicrobial activity against these bacteria and fungi than wild-type HSP-1, and therefore the prepared pharmaceutical composition should also have stronger antimicrobial properties against these bacteria and fungi.

[0050] Based on a general inventive concept, an embodiment of the present application provides an antibacterial drug, which includes the active ingredient of the antimicrobial peptide HSP-M4.

[0051] The antibacterial drug is realized based on the antibacterial peptide HSP-M4. The specific amino acid sequence of the antibacterial peptide HSP-M4 can refer to the above embodiments. Since the antibacterial drug adopts part or all of the technical solutions of the above embodiments, it has at least all the beneficial effects brought by the technical solutions of the above embodiments, which will not be described one by one here.

[0052] In some optional embodiments, the antibacterial drug is a drug for treating bacterial infection and / or fungal infection.

[0053] In the examples of the present application, the specific classification of antibacterial drugs is limited because the antibacterial drugs contain the active ingredient of the designed antimicrobial peptide HSP-M4, and thus are drugs that can effectively treat bacterial or fungal infections.

[0054] In some optional embodiments, the bacterial infection is caused by Escherichia coli and / or Staphylococcus aureus; and / or,

[0055] The fungal infection is caused by Candida albicans.

[0056] In the examples of the present application, specific types of bacterial infection and fungal infection are defined. Since the designed antimicrobial peptide HSP-M4 has more effective antimicrobial activity against these bacteria and fungi than wild-type HSP-1, the antimicrobial drug containing the antimicrobial peptide HSP-M4 should also have stronger antimicrobial properties against these bacteria and fungi.

[0057] The present application will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are intended only to illustrate the present application and are not intended to limit the scope of the present application. The experimental methods for which specific conditions are not specified in the following examples are usually measured according to national standards. If there is no corresponding national standard, then the conditions recommended by the manufacturer are followed.

[0058] Example 1

[0059] Rational molecular design of HSP series derivative peptides:

[0060] HSP-1 has low hemolytic and cytotoxic activity and can be used as a candidate compound for the development of antibacterial drugs. Therefore, the wild-type HSP-1 target sequence was used as the starting point for optimization. The optimization results are shown in Table 1.

[0061] Table 1 Parameters of optimized helical structure of wild-type HSP-1

[0062]

[0063] In Table 1, the bold parts are optimized amino acid sites.

[0064] Since the net charge and hydrophobicity of the peptide have a certain influence on the antimicrobial peptide activity, Figure 3 As shown in Table 1, the third amino acid D in the HSP-1 sequence was mutated to A to obtain HSP-M1, whose amino acid sequence is shown in SEQ ID NO.2, and the net charge number is increased to z=2.

[0065] To improve the hydrophobic properties (H) of HSP-1 and increase the hydrophobic distance (μH), combined with online Schiffie-Edumundson prediction analysis (heliQuest software analysis: https: / / heliquest.ipmc.cnrs.fr / ), e.g. Figure 3 As shown in Table 1, the hydrophobic amino acid L was used to replace the A in the hydrophobic surface to obtain HSP-M2, whose amino acid sequence is shown in SEQ ID NO.3.

[0066] To further increase the net charge of antimicrobial peptides, Figure 3 As shown in Table 1, the number of lysine in the sequence was increased to obtain HSP-M3 and HSP-M4 (with net charge numbers of 2 and 4, respectively), whose amino acid sequences are shown in SEQ ID NO.4 and SEQ ID NO.1, respectively; Figure 3 As shown in Table 1, HSP-M5 was obtained by increasing the charge and hydrophobicity on the basis of HSP-M4, and its amino acid sequence is shown in SEQ ID NO.5.

[0067] Many studies have shown that the helical structure of antimicrobial peptides is crucial for their antimicrobial activity. Figure 3 As shown in Table 1, the N-terminal of the HSP-1 sequence was truncated by 4 amino acids (equivalent to an α-helical structure) to obtain HSP-M6, whose amino acid sequence is shown in SEQ ID NO.6, and the C-terminal was truncated by 4 amino acids to obtain HSP-M7, whose amino acid sequence is shown in SEQ ID NO.7, in order to facilitate the investigation of the effect of the shortened helical structure on its antibacterial activity.

[0068] Similarly, whether the extension of the α-helix will affect its antibacterial activity also needs to be investigated. Four amino acids were added from the N-terminus and C-terminus of HSP-1 to extend the α-helix, and the added amino acids were mainly glycine and alanine that are easy to form a helical structure (such as Figure 1 and HSP-M8 and HSP-M9 in Table 1, whose amino acid sequences are shown in SEQ ID NO.8 and SEQ ID NO.9, respectively).

[0069] like Figure 4 As shown in Figure 2, the secondary structure of HSP-derived peptides with truncated and increased α-helical structures was predicted (online software I-TASSER prediction: https: / / zhanggroup.org / ). The secondary structure showed that compared with wild-type HSP-1, the α-helices in the secondary structures of truncated HSP-M6 and HSP-M7 were significantly reduced, but the α-helices in the N-terminal truncated HSP-M6 were less than those in the C-terminal truncated HSP-M7 (e.g. Figure 4 After adding α-helical sequences to the N-terminus and C-terminus of HSP-1, the antimicrobial peptides HSP-M8 and HSP-M9 have correspondingly increased 1-2 α-helical structures (as shown in Figure 4 Its antibacterial activity needs to be further demonstrated.

[0070] Example 2

[0071] Comparing Example 2 with Example 1, the difference between Example 2 and Example 1 is:

[0072] Antimicrobial activity testing of HSP series derived peptides:

[0073] Antibacterial activity test: S. coli ATCC 25922 and S. aureus ATCC 29213 were inoculated in Mueller-Hinton liquid medium (MH medium) at 37°C and cultured in a shaking incubator at 220 rpm. When the cells reached the logarithmic phase, MH medium was used as the diluent to dilute the cells appropriately and transfer to a 96-well plate. 90 μL was inoculated into each well to make the final concentration of the cells reach 5×10 5 cfu / mL.

[0074] MH medium was configured with HSP series derivative peptides, with the highest final concentration of 256μg / mL. 12 concentration gradients were formed by dilution, and 10μL was inoculated in each well. The well without peptide drug was used as the control well. The final volume of each well of the 96-well plate was 100μL, and three wells were repeated for each concentration. After static culture at 7℃ for 12h, the absorbance was detected at 595nm on the microplate reader. Compared with the control well, the drug concentration in the lowest concentration well with an OD value of less than 80% was the minimum inhibitory concentration (MIC), and the results are shown in Table 2.

[0075] Antifungal activity test: Candida albicans ATCC 90029 was inoculated in Sabouraud dextrose broth (SDB) medium and cultured in a shaking incubator at 33°C and 220 rpm for 24 h. The bacterial solution was resuspended in 5 mL of sterile saline and diluted with RPMI-1640 medium to a bacterial concentration of 0.5 × 10 3 ~2.5×10 3 CFU / mL, inoculate 90μL in a 96-well plate. 1640 medium is configured with HSP series derivative peptides, with the highest final concentration of 256μg / mL, and 12 concentration gradients are formed by double dilution, and 10μL is inoculated in each well. The well without peptide drug is used as the control well. The final volume of each well of the 96-well plate is 100μL, and each concentration is repeated three times. After static culture at 33℃ for 24h, the absorbance is detected at 595nm on an enzyme reader. Compared with the control well, the drug concentration in the lowest concentration well where the OD value drops by less than 80% is the MIC value. The results are shown in Table 2.

[0076] Table 2 Antimicrobial activity test results of various antimicrobial peptides

[0077]

[0078] As shown in Table 2, the MIC of HSP-M4 against E. coli ATCC 25922 and S. aureus ATCC 29213 were both 2 μg / mL, which were 64 and 32 times higher than that of natural HSP-1, respectively.

[0079] As shown in Table 2, the MIC of HSP-M4 against C.albicans ATCC 90029 is 64 μg / mL, which is 4 times higher than that of natural HSP-1.

[0080] Example 3

[0081] Comparing Example 3 with Example 1, the difference between Example 3 and Example 1 is:

[0082] Cytotoxicity test of HSP series derived peptides:

[0083] Hemolysis experiment: Fresh blood (containing anticoagulant) collected from the hospital was transferred to a sterile 10mL centrifuge tube, centrifuged at 210×g for 10 min, the supernatant was removed and the red blood cell pellet was retained, cold PBS was added and gently mixed, and then centrifuged again. This step was repeated until the supernatant was clear after centrifugation.

[0084] Finally, the supernatant was removed to prepare packed red blood cells, and the red blood cells were diluted with PBS to prepare a 2% red blood cell suspension. The maximum final concentration of HSP polypeptide was 256μM, and 8 concentration gradients were diluted by fold, and 100μL was added to each centrifuge tube; 100μL of 2% red blood cell suspension was added to each centrifuge tube, and gently mixed. Triton X-100 with a final concentration of 0.1% was added to the positive control well, and PBS was added to the negative control. Each well was repeated three times and incubated in a 37℃ incubator for 1h. Centrifuge at 1000×g for 20min, and 100μL of supernatant was transferred from each centrifuge tube to 96 wells, and the absorbance value was measured at 576nm.

[0085] The hemolysis rate was calculated according to the following formula: Hemolysis rate % = (OD576nm sample - OD576nm negative) / (OD576nm positive - OD576nm negative) × 100%. Figure 5 shown.

[0086] from Figure 5 It can be seen that the erythrocyte hemolysis rate of HSP-M4 is less than 10%, indicating that HSP-M4 has low toxicity to human erythrocytes.

[0087] MTT cytotoxicity assay: 90 μL of HEK293 cells in the proliferation stage were inoculated into a 96-well plate at a concentration of 5 × 10 4 cells, and cultured in a 37°C carbon dioxide incubator for 6 hours. Then add 10μL of HSP peptide drugs, the final concentration of peptide drugs is 128μM, 64μM, 32μM, 16μM, 8μM, 4μM, 2μM, 1μM, and cultured in a 37°C carbon dioxide incubator for 24 hours. After the incubation, add 10μL of MTT to each well for 4 hours, then add 100μL of DMSO to each well to dissolve, and detect the absorbance at a wavelength of 570nm. No cells are added to the blank group, only culture medium is added; no drug is added to the control group, cells and culture medium are added, and each well is repeated three times.

[0088] The cell survival rate was calculated according to the following formula: Cell survival rate % = (OD570nm sample - OD570nm blank group) / (OD570nm control group - OD570nm blank group) × 100%. Figure 6 shown.

[0089] from Figure 6 It can be seen that under the action of HSP-M4, the survival rate of HEK293 cells is higher than 90%, indicating that HSP-M4 has low toxicity to human HEK293 cells.

[0090] In summary, a series of HSP derivative peptides were obtained by molecular modification of the existing natural antimicrobial peptide HSP-1. Antimicrobial activity detection showed that the antimicrobial activity of HSP-M4, a derivative peptide of the antimicrobial peptide HSP-1, was greatly improved. Its minimum inhibitory concentration (MIC) against Escherichia coli E. coli ATCC 25922 and Staphylococcus aureus S. aureus ATCC 29213 was 2 μg / mL, and its MIC against Candida albicans C. albicans ATCC 90029 was 64 μg / mL. At the same time, cytotoxicity tests showed that HSP-M4 had low toxic and side effects. At the same time, the present application provides a new candidate molecule for the development of innovative antimicrobial drugs.

[0091] Various embodiments of the present application may be presented in the form of a range; it should be understood that the description in the form of a range is only for convenience and brevity, and should not be understood as a rigid limitation on the scope of the present application; therefore, the range description should be considered to have specifically disclosed all possible sub-ranges and single numerical values ​​within the range. For example, the range description from 1 to 6 should be considered to have specifically disclosed sub-ranges, such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., as well as single numbers within the range, such as 1, 2, 3, 4, 5 and 6, which apply regardless of the range. In addition, whenever a numerical range is indicated herein, it is meant to include any cited number (fractional or integer) within the indicated range.

[0092] In the present application, in the absence of any contrary description, the directional words used, such as "upper" and "lower", are specifically the directions of the drawings in the accompanying drawings. In addition, in the description of the present specification, the terms "including", "comprising", etc. refer to "including but not limited to". In this article, relational terms such as "first" and "second" are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply that there is any such actual relationship or order between these entities or operations. In this article, "and / or" describes the association relationship of the associated objects, indicating that there may be three relationships, for example, A and / or B, which can represent: A exists alone, A and B exist at the same time, and B exists alone. Wherein A, B can be singular or plural. In this article, "at least one" refers to one or more, and "plural" refers to two or more. "At least one", "at least one of the following" or similar expressions refer to any combination of these items, including any combination of singular or plural items. For example, "at least one of a, b, or c", or "at least one of a, b and c", can both mean: a, b, c, ab (i.e. a and b), ac, bc or abc, where a, b, c can be single or plural, respectively.

[0093] The above description is only a specific implementation of the present application, so that those skilled in the art can understand or implement the present application. Various modifications to these embodiments will be apparent to those skilled in the art, and the general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present application. Therefore, the present application will not be limited to the embodiments shown herein, but will conform to the widest range consistent with the principles and novel features applied for herein.

Claims

1. An antimicrobial peptide HSP-M4 constructed based on rational molecular design, characterized in that: The amino acid sequence of the antimicrobial peptide HSP-M4 is shown in SEQ ID NO.

1. The antimicrobial peptide HSP-M4 is a derivative peptide of the antimicrobial peptide HSP-1.

2. The antimicrobial peptide HSP-M4 according to claim 1, characterized in that The C-terminus of the antimicrobial peptide HSP-M4 is an acetamidate terminal.

3. Use of the antimicrobial peptide HSP-M4 as claimed in claim 1 or 2 in the preparation of a pharmaceutical composition, wherein the pharmaceutical composition is used to fight against Escherichia coli, Stephila aureus and Candida albicans.

4. An antibacterial drug, characterized in that: The antibacterial drug comprises the active ingredient of the antibacterial peptide HSP-M4 as claimed in claim 1 or 2.

5. The antibacterial drug according to claim 4, characterized in that: The antibacterial drug is a drug for treating bacterial infection and / or fungal infection.

6. The antibacterial drug according to claim 4, characterized in that: The bacterial infection is caused by Escherichia coli and / or Staphylococcus aureus; and / or, The fungal infection is caused by Candida albicans.

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