CN114516899B - 抗菌肽yhx-5及其应用 - Google Patents
抗菌肽yhx-5及其应用 Download PDFInfo
- Publication number
- CN114516899B CN114516899B CN202210102034.6A CN202210102034A CN114516899B CN 114516899 B CN114516899 B CN 114516899B CN 202210102034 A CN202210102034 A CN 202210102034A CN 114516899 B CN114516899 B CN 114516899B
- Authority
- CN
- China
- Prior art keywords
- yhx
- antibacterial peptide
- antibacterial
- peptide
- leu
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 239000003910 polypeptide antibiotic agent Substances 0.000 title claims abstract description 64
- 230000000844 anti-bacterial effect Effects 0.000 claims abstract description 24
- 241000588724 Escherichia coli Species 0.000 claims abstract description 10
- 241000186779 Listeria monocytogenes Species 0.000 claims abstract description 10
- 241000607142 Salmonella Species 0.000 claims abstract description 10
- 241000194019 Streptococcus mutans Species 0.000 claims abstract description 10
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract 2
- 239000003242 anti bacterial agent Substances 0.000 claims description 11
- 230000002401 inhibitory effect Effects 0.000 claims description 11
- 239000003755 preservative agent Substances 0.000 claims description 10
- 230000002335 preservative effect Effects 0.000 claims description 9
- 108700042778 Antimicrobial Peptides Proteins 0.000 claims description 8
- 102000044503 Antimicrobial Peptides Human genes 0.000 claims description 8
- 239000003599 detergent Substances 0.000 claims description 8
- 239000000203 mixture Substances 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 4
- 241001465754 Metazoa Species 0.000 claims description 3
- -1 body wash Substances 0.000 claims description 2
- 239000002537 cosmetic Substances 0.000 claims description 2
- 235000013305 food Nutrition 0.000 claims description 2
- 239000000843 powder Substances 0.000 claims description 2
- 238000002360 preparation method Methods 0.000 claims description 2
- 239000002453 shampoo Substances 0.000 claims description 2
- 239000000344 soap Substances 0.000 claims description 2
- 229940034610 toothpaste Drugs 0.000 claims description 2
- 239000000606 toothpaste Substances 0.000 claims description 2
- 230000002949 hemolytic effect Effects 0.000 abstract description 7
- 238000003786 synthesis reaction Methods 0.000 abstract description 7
- 150000001413 amino acids Chemical class 0.000 description 9
- 230000001580 bacterial effect Effects 0.000 description 8
- 108090000765 processed proteins & peptides Proteins 0.000 description 8
- 102000004196 processed proteins & peptides Human genes 0.000 description 7
- 238000004458 analytical method Methods 0.000 description 5
- 229940088710 antibiotic agent Drugs 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 239000001963 growth medium Substances 0.000 description 5
- 241000894006 Bacteria Species 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 210000000170 cell membrane Anatomy 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 230000007246 mechanism Effects 0.000 description 4
- 239000013642 negative control Substances 0.000 description 4
- 125000003345 AMP group Chemical group 0.000 description 3
- 206010059866 Drug resistance Diseases 0.000 description 3
- 241000192125 Firmicutes Species 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 230000004071 biological effect Effects 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 229920006227 ethylene-grafted-maleic anhydride Polymers 0.000 description 3
- 230000002209 hydrophobic effect Effects 0.000 description 3
- 230000003834 intracellular effect Effects 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- 238000004659 sterilization and disinfection Methods 0.000 description 3
- 206010018910 Haemolysis Diseases 0.000 description 2
- 241000191967 Staphylococcus aureus Species 0.000 description 2
- 244000052616 bacterial pathogen Species 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 238000009510 drug design Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 210000003743 erythrocyte Anatomy 0.000 description 2
- 230000008588 hemolysis Effects 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 108020004707 nucleic acids Proteins 0.000 description 2
- 102000039446 nucleic acids Human genes 0.000 description 2
- 150000007523 nucleic acids Chemical class 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 239000013049 sediment Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 101100379209 Arabidopsis thaliana APD3 gene Proteins 0.000 description 1
- 108010077805 Bacterial Proteins Proteins 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 108010026389 Gramicidin Proteins 0.000 description 1
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108010053775 Nisin Proteins 0.000 description 1
- NVNLLIYOARQCIX-MSHCCFNRSA-N Nisin Chemical compound N1C(=O)[C@@H](CC(C)C)NC(=O)C(=C)NC(=O)[C@@H]([C@H](C)CC)NC(=O)[C@@H](NC(=O)C(=C/C)/NC(=O)[C@H](N)[C@H](C)CC)CSC[C@@H]1C(=O)N[C@@H]1C(=O)N2CCC[C@@H]2C(=O)NCC(=O)N[C@@H](C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(NCC(=O)N[C@H](C)C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCSC)C(=O)NCC(=O)N[C@H](CS[C@@H]2C)C(=O)N[C@H](CC(N)=O)C(=O)N[C@H](CCSC)C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(N[C@H](C)C(=O)N[C@@H]3C(=O)N[C@@H](C(N[C@H](CC=4NC=NC=4)C(=O)N[C@H](CS[C@@H]3C)C(=O)N[C@H](CO)C(=O)N[C@H]([C@H](C)CC)C(=O)N[C@H](CC=3NC=NC=3)C(=O)N[C@H](C(C)C)C(=O)NC(=C)C(=O)N[C@H](CCCCN)C(O)=O)=O)CS[C@@H]2C)=O)=O)CS[C@@H]1C NVNLLIYOARQCIX-MSHCCFNRSA-N 0.000 description 1
- 108091005461 Nucleic proteins Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 108010040201 Polymyxins Proteins 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 125000001931 aliphatic group Chemical group 0.000 description 1
- 230000000843 anti-fungal effect Effects 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000010100 anticoagulation Effects 0.000 description 1
- 229940121375 antifungal agent Drugs 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 230000009286 beneficial 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
- 238000004364 calculation method Methods 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 229960004905 gramicidin Drugs 0.000 description 1
- ZWCXYZRRTRDGQE-SORVKSEFSA-N gramicidina Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CC=3C4=CC=CC=C4NC=3)NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CC=3C4=CC=CC=C4NC=3)NC(=O)[C@@H](CC(C)C)NC(=O)[C@H](CC=3C4=CC=CC=C4NC=3)NC(=O)[C@H](C(C)C)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](C(C)C)NC(=O)[C@H](C)NC(=O)[C@H](NC(=O)[C@H](C)NC(=O)CNC(=O)[C@@H](NC=O)C(C)C)CC(C)C)C(=O)NCCO)=CNC2=C1 ZWCXYZRRTRDGQE-SORVKSEFSA-N 0.000 description 1
- 229960002897 heparin Drugs 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- 238000000265 homogenisation Methods 0.000 description 1
- 230000005661 hydrophobic surface Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000011031 large-scale manufacturing process Methods 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000004309 nisin Substances 0.000 description 1
- 235000010297 nisin Nutrition 0.000 description 1
- 238000000424 optical density measurement Methods 0.000 description 1
- 230000000590 parasiticidal effect Effects 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000001243 protein synthesis Methods 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000003362 replicative effect Effects 0.000 description 1
- 150000003384 small molecules Chemical class 0.000 description 1
- 238000005556 structure-activity relationship Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 230000014616 translation Effects 0.000 description 1
- GPRLSGONYQIRFK-MNYXATJNSA-N triton Chemical compound [3H+] GPRLSGONYQIRFK-MNYXATJNSA-N 0.000 description 1
- 229910021642 ultra pure water Inorganic materials 0.000 description 1
- 239000012498 ultrapure water Substances 0.000 description 1
Images
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
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
- A01N37/44—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a nitrogen atom attached to the same carbon skeleton by a single or double bond, this nitrogen atom not being a member of a derivative or of a thio analogue of a carboxylic group, e.g. amino-carboxylic acids
- A01N37/46—N-acyl derivatives
-
- 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
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
- A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
- A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
- A23L3/3463—Organic compounds; Microorganisms; Enzymes
- A23L3/3526—Organic compounds containing nitrogen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
-
- 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
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q11/00—Preparations for care of the teeth, of the oral cavity or of dentures; Dentifrices, e.g. toothpastes; Mouth rinses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q17/00—Barrier preparations; Preparations brought into direct contact with the skin for affording protection against external influences, e.g. sunlight, X-rays or other harmful rays, corrosive materials, bacteria or insect stings
- A61Q17/005—Antimicrobial preparations
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/10—Washing or bathing preparations
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q5/00—Preparations for care of the hair
- A61Q5/02—Preparations for cleaning the hair
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/10—General cosmetic use
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
- A61K2800/52—Stabilizers
- A61K2800/524—Preservatives
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Polymers & Plastics (AREA)
- Organic Chemistry (AREA)
- Dermatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Food Science & Technology (AREA)
- Biophysics (AREA)
- Birds (AREA)
- Biochemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Oncology (AREA)
- Communicable Diseases (AREA)
- Animal Husbandry (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Epidemiology (AREA)
- Pharmacology & Pharmacy (AREA)
- Microbiology (AREA)
- Nutrition Science (AREA)
- Agronomy & Crop Science (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Environmental Sciences (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
本发明属于生物技术领域。针对现有的抗菌肽不能满足临床应用的上述问题，本发明提供一种抗菌肽YHX‑5及其应用。所述抗菌肽YHX‑5的氨基酸序列为：Gly‑Lys‑Leu‑Leu‑Ser‑Lys‑Leu‑Leu‑Gly‑Lys‑Leu‑Leu‑Lys‑NH2。本发明的抗菌肽对单核细胞增生李斯特氏菌、变异链球菌、大肠杆菌及沙门氏菌均有较好的抗菌活性。本发明的抗菌肽溶血活性非常低，安全性极高，合成序列短，化学合成难度小。
Description
技术领域
本发明属于生物技术领域，具体涉及一种抗菌肽及其应用。
背景技术
抗生素无疑是人类医药历史上最伟大的发现之一，已经拯救了数以千万人的生命。抗生素的抑菌机理主要包括四个方面：抑制细菌细胞壁的合成，增强细菌细胞膜的通透性，干扰细菌蛋白质合成以及抑制核酸的复制转录过程。但近年来抗生素滥用导致耐药菌株不断增多及多重耐药菌的出现，对治疗临床感染构成了严重的威胁。因此解决病原菌的耐药性问题已经刻不容缓。而开发新型抗菌药物是解决此问题的一个重要途径。其中抗菌肽凭借独特的杀菌机制以及快速的杀菌效率，不易产生耐药性，成为抗生素最有力的替代者。
抗菌肽(antimicrobial peptides，AMPs)是一类小分子活性多肽，一般由10-50个氨基酸组成。抗菌肽具有包括抗细菌、抗真菌、抗病毒、杀灭寄生虫、抑制肿瘤以及调节免疫系统的多种生物活性。抗菌肽的作用靶点具有多重性，既可直接作用于细胞膜，破坏细胞膜完整性，又可作用于细胞内靶点，进而抑制核酸、蛋白质的合成或者抑制胞内酶的活性。其独特的膜破坏机制和胞内杀菌机制不易产生耐药性，因而具有广泛的应用前景。
截止到2019年5月，权威的抗菌肽数据库APD3(http://aps.unmc.edu/AP/main.php)已经收录了3167条抗菌肽序列。但目前只有少数的抗菌肽被用于临床治疗，如乳酸链球菌肽、短杆菌肽和多粘菌素等。造成抗菌肽在实际应用中如此有限的原因主要是天然存在的抗菌肽有几项缺陷：(1)抗菌肽生物活性较低，达不到临床应用标准，无法从根本上替代抗生素；(2)溶血活性高；(3)生产成本高。因此，通过理性设计开发出结构简单、生物活性高同时无明显毒副作用的抗菌肽是当前亟需解决的问题。
发明内容
针对现有的抗菌肽不能满足临床应用的上述问题，本发明提供一种抗菌肽YHX-5及其应用，本发明的抗菌肽对单核细胞增生李斯特氏菌、变异链球菌、大肠杆菌及沙门氏菌均有较好的抗菌活性，并且溶血活性非常低，安全性极高，合成序列短，化学合成难度小。
为实现上述目的，本发明提供一种抗菌肽YHX-5，所述抗菌肽YHX-5的氨基酸序列为：Gly-Lys-Leu-Leu-Ser-Lys-Leu-Leu-Gly-Lys-Leu-Leu-Lys-NH2，在序列表中的位置为SEQ ID NO.1。
本发明的一个方面，提供所述抗菌肽YHX-5的抗菌用途。
进一步，所述抗菌用途为抗单核细胞增生李斯特氏菌、变异链球菌、沙门氏菌以及大肠杆菌。
进一步，所述抗菌肽YHX-5对单核细胞增生李斯特氏菌的最小抑菌浓度为128μg/mL。
进一步，所述抗菌肽YHX-5对变异链球菌的最小抑菌浓度为32μg/mL。
进一步，所述抗菌肽YHX-5对沙门氏菌的最小抑菌浓度为16μg/mL。
进一步，所述抗菌肽YHX-5对大肠杆菌的最小抑菌浓度为64μg/mL。
本发明还提供一种生物抗菌剂，其抗菌成分包含上述抗菌肽YHX-5。
本发明还提供一种动物饲料，其含有上述抗菌肽YHX-5。
本发明还提供一种防腐剂，其含有上述抗菌肽YHX-5。
优选地，所述防腐剂为用于食品的防腐剂或者用于化妆品的防腐剂。
本发明还提供一种洗涤剂组合物，其含有上述抗菌肽YHX-5。
优选地，所述洗涤剂组合物为洗手液、香皂、沐浴液、洗发剂、牙膏、洗衣液或洗衣粉等。
本发明的有益效果在于：
(1)本发明具有广谱的抗菌活性，能够抑制革兰氏阳性菌和革兰氏阴性菌。对革兰氏阳性菌例如单核细胞增生李斯特氏菌、变异链球菌等，革兰氏阴性菌例如沙门氏菌、大肠杆菌等，均具有较高的抗菌活性。
(2)本发明所述的抗菌肽，溶血活性非常低，合成序列短，分子量小，化学合成难度小，能够更加特异性地杀死生物体内的病原菌同时很好的节约规模化生产成本。
附图说明
图1为本发明实施例1提供的抗菌肽YHX-5的二级结构预测示意图；
图2为本发明实施例2提供的抗菌肽YHX-5在333μg/mL浓度条件下对单核细胞增生李斯特氏菌的抑菌效果示意图。
图3为本发明实施例2提供的抗菌肽YHX-5在333μg/mL浓度条件下对变异链球菌的抑菌效果示意图。
图4为本发明实施例2提供的抗菌肽YHX-5在333μg/mL浓度条件下对沙门氏菌的抑菌效果示意图。
图5为本发明实施例2提供的抗菌肽YHX-5在333μg/mL浓度条件下对大肠杆菌的抑菌效果示意图。
具体实施方式
下面结合具体实施例对本发明做进一步详细说明。
实施例1抗菌肽的设计
在抗菌肽从头设计及对构效关系理解的基础上，从ADP3数据库中筛选得到的对革兰氏阳性菌和革兰氏阴性菌有抑菌作用的843条抗菌肽序列进行序列参数分析(包括序列长度、带电荷数、疏水氨基酸比例和氨基酸组成)，之后按照优势参数的选取原则并结合理性设计思路，确定了新抗菌肽的各项序列参数，具体如下：
表1抗菌肽序列的设计参数
选择在数据库中参数频数第二位的13作为新抗菌肽的序列长度，在降低分子量提高安全性的同时又可以有效降低合成成本。带电荷数+4在数据库参数频数中同样处于第一位，可以保证抗菌肽对细菌细胞膜的亲和力。选取带正电氨基酸Lys和极性不带电荷氨基酸Ser组成抗菌肽的极性面，选取疏水性氨基酸Leu形成疏水面，来保证抗菌肽的两亲性结构。由于Gly在各氨基酸占比中排列于第一位，因此在序列中引入两个Gly残基以稳定结构，同时Gly位于N-末端有利于提高抗菌肽半衰期，提高其应用潜力。
采用生物信息学预测及分析工具，对不同氨基酸排列顺序的肽序列分子量、平均疏水值、平均疏水力矩、不稳定性指数、脂肪族指数、半衰期、螺旋轮图、成为AMPs的可能性以及二级结构进行预测，选出最优的抗菌肽序列。所用生物信息学工具如下：
物化性质分析网站：Expasy ProtParam(https://web.expasy.org/protparam/)
螺旋轮图及疏水性分析网站：Heliquest(https://heliquest.ipmc.cnrs.fr/)
AMPs可能性预测网站：CAMPR3(http://www.camp.bicnirrh.res.in/)
二级结构预测网站：Pep-fold(https://bioserv.rpbs.univ-paris-diderot.fr/services/PEP-FOLD3/)；
Zhang Lab(https://zhanglab.ccmb.med.umich.edu/)
通过上述工具得到的最优抗菌肽序列及物化性质如表2所示，二级结构预测结果如图1所示。
表2抗菌肽YHX-5的序列及物化性质
实施例2抑菌活性
抗菌肽由强耀生物科技(上海)有限公司合成。
将金黄色葡萄球菌、大肠杆菌和沙门氏菌分别划线接种于LB固体培养基，将单核细胞增生李斯特氏菌和变异链球菌划线接种于BHI固体培养基，置于37℃恒温培养箱培养18h，挑取各个菌株的单菌落置于各自对应的液体培养基中，37℃恒温振荡培养12h。测量菌液的OD600值，并将其稀释至1×106CFU/mL。
①抑菌圈实验
配置LB和BHI半固体培养基(琼脂质量分数0.6％)，每皿20mL加入7μL菌液振荡混匀后倒入摆放好牛津杯的培养皿中，待培养基冷却凝固后拔除牛津杯完成打孔。每孔中加入160μL抗菌肽溶液(配置使用1mg抗菌肽溶于3mL超纯水)。置于37℃恒温培养箱培养24h后测量抑菌圈直径，测试结果如图2-5所示。
②最小抑菌浓度(MIC)的测定
向96孔板中加入培养至对数生长期的各菌株菌液(单核细胞增生李斯特氏菌、变异链球菌、金黄色葡萄球菌、沙门氏菌、大肠杆菌)50μL，同时在各孔中加入各浓度梯度抗菌肽溶液(512μg/mL、256μg/mL、128μg/mL、64μg/mL、32μg/mL、16μg/mL、8μg/mL、4μg/mL)50μL，并分别做阳性对照和阴性对照组。在37℃恒温培养箱培养8h后测量各孔的OD600值，取菌株生长被完全抑制的最低浓度作为抗菌肽的最小抑菌浓度(MIC)，测试结果如表3所示。
表3抗菌肽YHX-5的抑菌活性分析结果
实施例3溶血活性
取新鲜健康兔血液1mL加于肝素抗凝管中，在1000xg转速下离心10min后取沉淀，用PBS缓冲液冲洗3次，将红细胞用10mLPBS重悬。用PBS缓冲液将抗菌肽YHX-5的浓度调整至4-512μg/mL，以每孔50μL的剂量加入96孔板中，并加入等体积(50μL)的红细胞悬液混匀。以PBS缓冲液作为阴性对照，以0.1％Tritonx-100作为阴性对照，在37℃恒温培养1h后取出，1000xg离心10min，取出上清液用酶标仪在570nm处测OD值，测试结果如表4所示。
溶血率计算公式为：溶血率＝(AT-AO)/(AC-AO)×100％。
式中：AT为实验组的吸光值，AC为阳性对照组的吸光值，AO为阴性对照组的吸光值。
表4抗菌肽YHX-5的溶血活性(测定3次取平均值)
由表4可知，即使在512μg/mL的最大浓度下，抗菌肽YHX-5也未呈现明显的溶血活性，说明抗菌肽YHX-5有极好的安全性，在制作生物抗菌剂、动物饲料以及防腐剂等领域的应用前景较大，可进一步深入研究和开发利用。
以上所述的实施例仅是对本发明的优选实施方式进行描述，并非对本发明的范围进行限定，在不脱离本发明设计精神的前提下，本领域普通技术人员对本发明的技术方案作出的各种变形和改进，均应落入本发明权利要求书确定的保护范围内。
序列表
<110> 中国海洋大学
<120> 抗菌肽YHX-5及其应用
<141> 2022-01-26
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 13
<212> PRT
<213> Artificial Sequence
<400> 1
Gly Lys Leu Leu Ser Lys Leu Leu Gly Lys Leu Leu Lys
1 5 10
Claims (9)
1.抗菌肽YHX-5，其特征在于，所述的抗菌肽YHX-5的氨基酸序列如SEQ ID NO.1所示。
2.权利要求1所述的抗菌肽YHX-5在制备抑制单核细胞增生李斯特氏菌、变异链球菌、沙门氏菌以及大肠杆菌的生物抗菌剂中的应用。
3.根据权利要求2所述的应用，其特征在于，所述抗菌肽YHX-5对单核细胞增生李斯特氏菌的最小抑菌浓度为128μg/mL；对变异链球菌的最小抑菌浓度为32μg/mL；对沙门氏菌的最小抑菌浓度为16μg/mL；对大肠杆菌的最小抑菌浓度为64μg/mL。
4.一种生物抗菌剂，其特征在于，其抗菌成分中包含权利要求1所述的抗菌肽YHX-5。
5.一种动物饲料，其特征在于，含有如权利要求1所述的抗菌肽YHX-5。
6.一种防腐剂，其特征在于，含有如权利要求1所述的抗菌肽YHX-5。
7.根据权利要求6所述的防腐剂，其特征在于，所述防腐剂为用于食品的防腐剂或者用于化妆品的防腐剂。
8.一种洗涤剂组合物，其特征在于，含有如权利要求1所述的抗菌肽YHX-5。
9.根据权利要求8所述的洗涤剂组合物，其特征在于，所述洗涤剂组合物为洗手液、香皂、沐浴液、洗发剂、牙膏、洗衣液或洗衣粉。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210102034.6A CN114516899B (zh) | 2022-01-27 | 2022-01-27 | 抗菌肽yhx-5及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210102034.6A CN114516899B (zh) | 2022-01-27 | 2022-01-27 | 抗菌肽yhx-5及其应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114516899A CN114516899A (zh) | 2022-05-20 |
| CN114516899B true CN114516899B (zh) | 2023-06-16 |
Family
ID=81596317
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210102034.6A Active CN114516899B (zh) | 2022-01-27 | 2022-01-27 | 抗菌肽yhx-5及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114516899B (zh) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115724909B (zh) * | 2022-08-29 | 2023-11-28 | 扬州市扬大康源乳业有限公司 | 抗菌肽yhx-7及其应用 |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017222335A1 (ko) * | 2016-06-23 | 2017-12-28 | 건국대학교 산학협력단 | 그람음성 다제내성균에 대해 항생제와 상승적 항균 효과를 가지는 항생 펩타이드 및 이의 용도 |
| WO2018086516A1 (zh) * | 2016-11-09 | 2018-05-17 | 湖南科技学院 | 一种抑制耐药性细菌的抗菌肽及其应用 |
| CN111944020A (zh) * | 2020-08-18 | 2020-11-17 | 中国海洋大学 | 抗菌肽及其应用 |
| CN112940082A (zh) * | 2021-04-23 | 2021-06-11 | 扬州市扬大康源乳业有限公司 | 一种抗菌肽及其应用 |
| CN113336827A (zh) * | 2021-06-30 | 2021-09-03 | 中国海洋大学 | 一种低分子量抗菌肽yhx-2及其组合物和应用 |
| CN113336828A (zh) * | 2021-06-30 | 2021-09-03 | 中国海洋大学 | 抗菌肽yhx-3及其组合物和应用 |
-
2022
- 2022-01-27 CN CN202210102034.6A patent/CN114516899B/zh active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017222335A1 (ko) * | 2016-06-23 | 2017-12-28 | 건국대학교 산학협력단 | 그람음성 다제내성균에 대해 항생제와 상승적 항균 효과를 가지는 항생 펩타이드 및 이의 용도 |
| WO2018086516A1 (zh) * | 2016-11-09 | 2018-05-17 | 湖南科技学院 | 一种抑制耐药性细菌的抗菌肽及其应用 |
| CN111944020A (zh) * | 2020-08-18 | 2020-11-17 | 中国海洋大学 | 抗菌肽及其应用 |
| CN112940082A (zh) * | 2021-04-23 | 2021-06-11 | 扬州市扬大康源乳业有限公司 | 一种抗菌肽及其应用 |
| CN113336827A (zh) * | 2021-06-30 | 2021-09-03 | 中国海洋大学 | 一种低分子量抗菌肽yhx-2及其组合物和应用 |
| CN113336828A (zh) * | 2021-06-30 | 2021-09-03 | 中国海洋大学 | 抗菌肽yhx-3及其组合物和应用 |
Non-Patent Citations (1)
| Title |
|---|
| 产抗菌肽乳酸菌筛选及抗菌肽的分离纯化与特性研究;章检明等;《食品安全质量检测学报》;第9卷(第4期);第781-787页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114516899A (zh) | 2022-05-20 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN112940082B (zh) | 一种抗菌肽及其应用 | |
| CN113336828B (zh) | 抗菌肽yhx-3及其组合物和应用 | |
| CN113336827B (zh) | 一种低分子量抗菌肽yhx-2及其组合物和应用 | |
| CN114516900B (zh) | 抗菌肽yhx-4及其应用 | |
| CN104151415B (zh) | 一种天然抗菌肽Alligatorin4及其应用 | |
| Lee et al. | Fungicidal mechanisms of the antimicrobial peptide Bac8c | |
| CN113121666B (zh) | 抗菌肽Scybaumancin105-127及其应用 | |
| CN114516899B (zh) | 抗菌肽yhx-5及其应用 | |
| CN102702333A (zh) | 抗耐药性病原菌感染多肽及其用途 | |
| CN106543271A (zh) | 抗耐药性感染多肽Cbf‑14‑2及其用途 | |
| CN114716512A (zh) | 一种青蟹广谱抗菌肽Scyampcin44-63及其应用 | |
| CN111808278A (zh) | 支化抗菌聚氨基酸及其制备方法与应用 | |
| CN104177485A (zh) | 一种扬子鳄抗菌肽Alligatorin6及其应用 | |
| CN104163861B (zh) | 一种爬行动物抗菌肽Alligatorin5及其应用 | |
| CN112430262A (zh) | 一类抗真菌肽及其应用 | |
| CN115724909B (zh) | 抗菌肽yhx-7及其应用 | |
| CN109820835B (zh) | 复合型胞内金黄色葡萄球菌抗菌制剂及其制备和应用 | |
| CN112724198A (zh) | 一种抗耐甲氧西林金黄色葡萄球菌抗菌肽及其制备方法和应用 | |
| CN117903253A (zh) | 一种抗菌肽rv15及其应用 | |
| CN111378023B (zh) | 一种抗变异链球菌多肽及其应用 | |
| CN117069819B (zh) | 一种黑腹狼蛛抗菌肽lc-amp-i1及其应用 | |
| CN109748949B (zh) | 一种抗菌肽及其应用 | |
| CN105273073B (zh) | 一种海蛇改造体抗菌肽qha2及其制备方法和应用 | |
| CN118005731A (zh) | 一种细菌素突变体bm-1及其应用 | |
| CN117986324A (zh) | 一种细菌素突变体am-1及其应用 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |