CN115558031B - Fusion peptide, collagen compound with antibacterial effect and application thereof - Google Patents
Fusion peptide, collagen compound with antibacterial effect and application thereof Download PDFInfo
- Publication number
- CN115558031B CN115558031B CN202211546201.2A CN202211546201A CN115558031B CN 115558031 B CN115558031 B CN 115558031B CN 202211546201 A CN202211546201 A CN 202211546201A CN 115558031 B CN115558031 B CN 115558031B
- Authority
- CN
- China
- Prior art keywords
- collagen
- lyase
- plyss2
- peptide
- fusion peptide
- 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
- 229920001436 collagen Polymers 0.000 title claims abstract description 94
- 108010035532 Collagen Proteins 0.000 title claims abstract description 92
- 102000008186 Collagen Human genes 0.000 title claims abstract description 92
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 34
- 230000004927 fusion Effects 0.000 title claims abstract description 16
- 230000000844 anti-bacterial effect Effects 0.000 title abstract description 24
- 150000001875 compounds Chemical class 0.000 title abstract description 12
- 102000004317 Lyases Human genes 0.000 claims abstract description 32
- 108090000856 Lyases Proteins 0.000 claims abstract description 32
- 241000894006 Bacteria Species 0.000 claims abstract description 9
- 230000003385 bacteriostatic effect Effects 0.000 claims description 27
- 230000000694 effects Effects 0.000 claims description 8
- 241001515965 unidentified phage Species 0.000 claims description 7
- 239000013604 expression vector Substances 0.000 claims description 6
- 241000588724 Escherichia coli Species 0.000 claims description 5
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 4
- 238000000576 coating method Methods 0.000 claims description 3
- 239000003814 drug Substances 0.000 claims description 3
- 235000013305 food Nutrition 0.000 claims description 3
- 238000002156 mixing Methods 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 241000295644 Staphylococcaceae Species 0.000 claims description 2
- 230000008618 cell wall macromolecule catabolic process Effects 0.000 claims description 2
- 239000011248 coating agent Substances 0.000 claims description 2
- 239000002537 cosmetic Substances 0.000 claims description 2
- 239000000945 filler Substances 0.000 claims description 2
- 239000000575 pesticide Substances 0.000 claims description 2
- 230000005764 inhibitory process Effects 0.000 claims 2
- 230000000845 anti-microbial effect Effects 0.000 claims 1
- 239000004599 antimicrobial Substances 0.000 claims 1
- 239000000825 pharmaceutical preparation Substances 0.000 claims 1
- 229940127557 pharmaceutical product Drugs 0.000 claims 1
- 102000037865 fusion proteins Human genes 0.000 abstract description 22
- 108020001507 fusion proteins Proteins 0.000 abstract description 22
- 239000000203 mixture Substances 0.000 abstract description 15
- 230000002035 prolonged effect Effects 0.000 abstract description 2
- 238000013329 compounding Methods 0.000 abstract 1
- 241000191967 Staphylococcus aureus Species 0.000 description 24
- 102000004169 proteins and genes Human genes 0.000 description 24
- 108090000623 proteins and genes Proteins 0.000 description 24
- 239000003102 growth factor Substances 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- 239000003242 anti bacterial agent Substances 0.000 description 9
- 239000013642 negative control Substances 0.000 description 9
- 210000001519 tissue Anatomy 0.000 description 8
- 238000000034 method Methods 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 238000012360 testing method Methods 0.000 description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 230000003796 beauty Effects 0.000 description 6
- 229940088710 antibiotic agent Drugs 0.000 description 5
- 239000000872 buffer Substances 0.000 description 5
- 239000007853 buffer solution Substances 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 5
- 210000002421 cell wall Anatomy 0.000 description 5
- 101000925662 Enterobacteria phage PRD1 Endolysin Proteins 0.000 description 4
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 4
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 210000002744 extracellular matrix Anatomy 0.000 description 4
- 238000007747 plating Methods 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 239000012460 protein solution Substances 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 230000008439 repair process Effects 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 206010059866 Drug resistance Diseases 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 101000595923 Homo sapiens Placenta growth factor Proteins 0.000 description 3
- 238000007865 diluting Methods 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 230000012010 growth Effects 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 208000015181 infectious disease Diseases 0.000 description 3
- 102000004196 processed proteins & peptides Human genes 0.000 description 3
- 208000017520 skin disease Diseases 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 208000035143 Bacterial infection Diseases 0.000 description 2
- 108020004705 Codon Proteins 0.000 description 2
- 201000004624 Dermatitis Diseases 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 108010014251 Muramidase Proteins 0.000 description 2
- 102000016943 Muramidase Human genes 0.000 description 2
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 2
- 206010052428 Wound Diseases 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 208000022362 bacterial infectious disease Diseases 0.000 description 2
- 239000003899 bactericide agent Substances 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 230000000975 bioactive effect Effects 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 230000035876 healing Effects 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 239000004325 lysozyme Substances 0.000 description 2
- 229960000274 lysozyme Drugs 0.000 description 2
- 235000010335 lysozyme Nutrition 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000012567 medical material Substances 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- 230000017423 tissue regeneration Effects 0.000 description 2
- 239000013598 vector Substances 0.000 description 2
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 1
- 241000589291 Acinetobacter Species 0.000 description 1
- 102000016359 Fibronectins Human genes 0.000 description 1
- 108010067306 Fibronectins Proteins 0.000 description 1
- 101001001487 Homo sapiens Phosphatidylinositol-glycan biosynthesis class F protein Proteins 0.000 description 1
- 101000740205 Homo sapiens Sal-like protein 1 Proteins 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 101100151529 Methanosarcina acetivorans (strain ATCC 35395 / DSM 2834 / JCM 12185 / C2A) lysS1 gene Proteins 0.000 description 1
- MSFSPUZXLOGKHJ-UHFFFAOYSA-N Muraminsaeure Natural products OC(=O)C(C)OC1C(N)C(O)OC(CO)C1O MSFSPUZXLOGKHJ-UHFFFAOYSA-N 0.000 description 1
- 241000186367 Mycobacterium avium Species 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- 108010013639 Peptidoglycan Proteins 0.000 description 1
- 102100035194 Placenta growth factor Human genes 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 1
- 102100037204 Sal-like protein 1 Human genes 0.000 description 1
- 241000607720 Serratia Species 0.000 description 1
- 206010072170 Skin wound Diseases 0.000 description 1
- 229920002385 Sodium hyaluronate Polymers 0.000 description 1
- 206010043376 Tetanus Diseases 0.000 description 1
- RTAQQCXQSZGOHL-UHFFFAOYSA-N Titanium Chemical compound [Ti] RTAQQCXQSZGOHL-UHFFFAOYSA-N 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 230000037358 bacterial metabolism Effects 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 238000010170 biological method Methods 0.000 description 1
- 229920001222 biopolymer Polymers 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 210000002808 connective tissue Anatomy 0.000 description 1
- 238000005336 cracking Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000003467 diminishing effect Effects 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000023597 hemostasis Effects 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 101150087853 lysK gene Proteins 0.000 description 1
- 101150110767 lysS gene Proteins 0.000 description 1
- 230000002101 lytic effect Effects 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 238000002264 polyacrylamide gel electrophoresis Methods 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000003259 recombinant expression Methods 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 230000037394 skin elasticity Effects 0.000 description 1
- 206010040872 skin infection Diseases 0.000 description 1
- 239000003009 skin protective agent Substances 0.000 description 1
- 229940010747 sodium hyaluronate Drugs 0.000 description 1
- YWIVKILSMZOHHF-QJZPQSOGSA-N sodium;(2s,3s,4s,5r,6r)-6-[(2s,3r,4r,5s,6r)-3-acetamido-2-[(2s,3s,4r,5r,6r)-6-[(2r,3r,4r,5s,6r)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2- Chemical compound [Na+].CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 YWIVKILSMZOHHF-QJZPQSOGSA-N 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 229910052719 titanium Inorganic materials 0.000 description 1
- 239000010936 titanium Substances 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 230000037314 wound repair Effects 0.000 description 1
- 230000037303 wrinkles Effects 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
-
- 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
- A01N47/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid
- A01N47/40—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid the carbon atom having a double or triple bond to nitrogen, e.g. cyanates, cyanamides
- A01N47/42—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid the carbon atom having a double or triple bond to nitrogen, e.g. cyanates, cyanamides containing —N=CX2 groups, e.g. isothiourea
- A01N47/44—Guanidine; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P1/00—Disinfectants; Antimicrobial compounds or mixtures thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/51—Lyases (4)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
- A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
- A61K47/6435—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent the peptide or protein in the drug conjugate being a connective tissue peptide, e.g. collagen, fibronectin or gelatin
-
- 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
- A61K8/65—Collagen; Gelatin; Keratin; Derivatives or degradation products thereof
-
- 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
- A61K8/66—Enzymes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
- A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
- A61L15/22—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing macromolecular materials
- A61L15/32—Proteins, polypeptides; Degradation products or derivatives thereof, e.g. albumin, collagen, fibrin, gelatin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
- A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
- A61L15/22—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing macromolecular materials
- A61L15/32—Proteins, polypeptides; Degradation products or derivatives thereof, e.g. albumin, collagen, fibrin, gelatin
- A61L15/325—Collagen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
- A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
- A61L15/42—Use of materials characterised by their function or physical properties
- A61L15/46—Deodorants or malodour counteractants, e.g. to inhibit the formation of ammonia or bacteria
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/227—Other specific proteins or polypeptides not covered by A61L27/222, A61L27/225 or A61L27/24
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/24—Collagen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/54—Biologically active materials, e.g. therapeutic substances
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
-
- 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
- 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
- A61Q19/005—Preparations for sensitive 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/88—Lyases (4.)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
- A61L2300/252—Polypeptides, proteins, e.g. glycoproteins, lipoproteins, cytokines
- A61L2300/254—Enzymes, proenzymes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/404—Biocides, antimicrobial agents, antiseptic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/80—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a special chemical form
- A61L2300/802—Additives, excipients, e.g. cyclodextrins, fatty acids, surfactants
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2800/00—Nucleic acids vectors
- C12N2800/10—Plasmid DNA
- C12N2800/101—Plasmid DNA for bacteria
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/185—Escherichia
- C12R2001/19—Escherichia coli
-
- 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
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Epidemiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Dermatology (AREA)
- Wood Science & Technology (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Pharmacology & Pharmacy (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- General Engineering & Computer Science (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Plant Pathology (AREA)
- Materials Engineering (AREA)
- Biotechnology (AREA)
- Hematology (AREA)
- Biophysics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Biochemistry (AREA)
- Transplantation (AREA)
- Pest Control & Pesticides (AREA)
- Birds (AREA)
- Gastroenterology & Hepatology (AREA)
- Microbiology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Environmental Sciences (AREA)
- Oncology (AREA)
- Communicable Diseases (AREA)
- Immunology (AREA)
Abstract
The invention belongs to the technical field of biology, and relates to a fusion peptide, a collagen compound with antibacterial effect and application thereof. According to the invention, the collagen compound with antibacterial effect is obtained by compounding the lyase-collagen combined peptide fusion protein with the collagen, compared with a single lyase and collagen mixture, the stability of the antibacterial activity of the lyase is improved, a new antibacterial function is given to the collagen, the resistance of the collagen to bacteria is improved, and the service life is prolonged.
Description
Technical Field
The invention belongs to the technical field of biology, and relates to a collagen compound with a bacteriostatic effect, and an application and a production method thereof.
Background
Collagen, also called collagen, is a biopolymer, the main component in animal ruminal tissues, and is also a functional protein with the largest content and the widest distribution in mammals, and accounts for 25-30% of the total protein. The collagen is the main component of the extracellular matrix, has unique chemical and spatial structure, has low immunogenicity, biological cracking property, hemostasis and repair functions, and plays an important role in the fields of modern tissue engineering, medical and aesthetic treatment and the like.
Traditional collagen is obtained mainly from animal connective tissue. In recent years, certain progress has been made in the fields of bone repair, skin wound repair dressing, muscle repair, drug delivery, beauty treatment and the like, and the recombinant human-like collagen is a protein which is optimized according to the characteristics of human collagen by a synthetic biological method and has the characteristics similar to the collagen through recombinant expression. Compared with the collagen obtained by traditional extraction, the collagen has the advantages of processability, no virus hidden trouble, good water solubility, stable production process, low rejection reaction and the like.
However, conventionally extracted collagen or recombinant human-like collagen is a rich biological macromolecule, and is very vulnerable to the attack of microorganisms such as bacteria. Research shows that bacterial metabolism can produce various proteases, so that chemical and spatial structures of collagen are damaged, and the biological activity of the collagen is reduced. The antibacterial and antiseptic properties are the necessary prerequisite for high-value utilization of collagen as medical materials and skin care products. The external addition of an antibacterial agent has proven to be an effective method. However, most of the commonly used antibacterial agents have physiological toxicity, and in clinical application, after collagen is absorbed and degraded, the contained antibacterial agents still have activity, and the environment accumulation thereof can cause the generation of bacterial drug resistance, so that the development of a novel biological antibacterial preservative is urgently needed to ensure that the collagen has antibacterial efficacy without generating drug resistance.
Besides high-value medical materials, the collagen has a large application market in the industries of medical beauty and beauty, and beauty and skin care. It has the functions of moistening skin, resisting wrinkle, maintaining skin smoothness and elasticity, repairing skin, promoting tissue healing, etc. The skin is used as a barrier for human health, and consists of skin epidermal microorganisms, superficial tissues and cells, various secretions, microenvironment and the like to form an ecosystem, namely the so-called skin micro-ecology. When the skin micro-ecology is destroyed, a series of skin diseases such as infection and enrichment of harmful bacteria on the skin are caused. Staphylococci are the most common causative bacteria of human skin infections, and in addition, escherichia coli, pseudomonas aeruginosa, tetanus, mycobacterium avium, and the like, and also, less commonly, acinetobacter, serratia, and the like, can cause various inflammations of the skin, such as: at present, the skin inflammation is commonly treated by using antibiotics or chemical bactericides and various skin protective agents (sodium hyaluronate, collagen, trehalose and the like) for assisting in treatment, but the antibiotics and the bactericides have broad spectrum and damage beneficial bacteria on the surface of the skin, and the use of the antibiotics can cause the appearance of drug-resistant strains, so that the stability of skin microecology is unfavorable, and an alternative treatment scheme is urgently needed.
In order to overcome the problem, a novel biological antibacterial agent, such as lysozyme, cell wall lyase derived from bacteriophage becomes an alternative method, and unlike the traditional antibiotics, lysozyme and the lyase derived from bacteriophage are both hydrolytic enzymes acting on bacterial cell walls, destroy the peptidoglycan structure of the bacterial cell walls, and are mild and free from drug resistance; moreover, most of the lytic enzymes derived from bacteriophages have a high specificity for the genus or species they infect, which has a significant advantage over classical broad-spectrum antibiotics.
How to enable the collagen to have the effects of moisturizing and maintaining the skin elasticity and the effects of resisting, inhibiting bacteria and diminishing inflammation of lyase in the using process, the collagen compound with the antibacterial effect is developed, the skin repair is accelerated, and the skin inflammation is prevented, so that the collagen compound has a potential application market.
In 2014 Mika ë lM. Martino et al (M.M. Martino, P.S. Briquez, E. Guc, F. Totelli and W.W. Kilarski et al, growth Factors Engineered for Super-Affinity to the Extracellular Matrix engineering titanium Healing, in Science, vol. 343, num 6173, p.885-888, 2014.) found a series of Extracellular Matrix ECM (including collagen, fibronectin etc.) binding peptides as follows: PIGF-2, etc., which researchers have conjugated to Growth Factor (GFs). The growth factor plays a key role in tissue repair, the extracellular matrix ECM exists in a large amount on a tissue affected part, the binding peptide increases the affinity between the growth factor and the tissue affected part, the growth factor is quickly aggregated and bound on a wound surface, the chronic wound tissue repair is accelerated, and the effect is obviously enhanced compared with that of singly using the growth factor GFs for treatment. In 2018, researchers of Shin-Hye Park et al ("All-in-one" in vitro selection of collagen-binding polypeptide endogenous growth factor. Biomaterials 161 (2018) 270-278) used growth factors to screen out various collagen-binding peptides, such as: SP1, SP2, etc., for localizing growth factors at the damaged site and enhancing the therapeutic effects of the growth factors.
In conclusion, the collagen-binding peptide is utilized to enable the lyase and the collagen to form a compound, so that the stability of the collagen can be improved, the resistance of the collagen to bacteria can be improved, the service life of the collagen can be prolonged, and a new antibacterial function of the collagen can be given, thereby being very beneficial to reducing the prevention and treatment cost, maintaining the skin micro-ecological balance, treating skin diseases and reducing the risk of bacterial infection of the affected part in the treatment process of the collagen product.
Disclosure of Invention
Based on the background, the invention combines cell wall lyase with antibacterial activity with collagen through the collagen binding peptide to obtain a collagen compound with an antibacterial function, thereby improving the stability of the cell wall lyase, reducing the risk of bacteria infection of the collagen, maintaining the stability of the collagen, prolonging the service life of the collagen, widening the application scene of the collagen on the basis of the original functions of the collagen and controlling the infection risk.
The invention provides a fusion peptide, which is formed by fusing lyase with a cell wall degradation effect and a collagen binding peptide.
Preferably, the lytic enzyme is derived from a bacteriophage.
More preferably, the lytic enzyme is bacteriophage lytic enzyme PlySs2, SAL-1, lysK, lysH5, phiII, MV-L, wherein the lytic enzyme PlySs2 has NCBI accession number WP _253220091.1.
Further preferably, the collagen binding peptide is the collagen binding peptide PIGF2.
The invention provides an expression vector containing the fusion peptide.
The invention also provides a recombinant strain containing the expression vector of the fusion peptide. Preferably, it is Escherichia coli.
The invention provides application of the fusion peptide in preparation of bacteriostatic products.
The invention particularly provides a collagen compound with antibacterial effect, which is characterized in that the collagen compound is obtained by mixing the fusion peptide with collagen.
The invention further provides application of the collagen compound with the bacteriostatic effect in preparing bacteriostatic products.
Specifically, the product refers to medicines, cosmetics, washing and caring products, medical dressings, medical tissue fillers, foods, food packages, antibacterial coatings and pesticides.
The invention provides a technical idea that new functions can be endowed to collagen through collagen binding peptide. Experiments show that the constructed lyase and collagen combined peptide fusion protein has a strong bacteriostatic action on staphylococcus aureus, is combined with collagen to endow the collagen with a new anti-bacteriostatic function, can prolong the service life of the collagen, and can prevent the collagen from being polluted. The constructed fusion protein is combined with collagen by utilizing collagen binding peptide PIGF to form a complex, so that the stability of the lyase is improved, and the bacteriostatic effect of the fusion protein is good when the fusion protein is directly mixed with the single lyase and the collagen. The complex has stable antibacterial effect, has practical application value, can be applied to the industries of modern tissue engineering, medical beauty, beauty treatment, skin care and the like, reduces the risk of bacterial infection of an affected part in the use process of a collagen product, and can maintain the microecological balance of skin to treat skin diseases and the like.
Drawings
FIG. 1: SDS-PAGE results of the purification of the PlsSs2 protein and PlsSs2-PIGF fusion protein. Wherein, 1 is PlsS 2 protein, and 2 is PlsS 2-PIGF.
FIG. 2: and (5) bacteriostatic effect graphs of different proteins. Wherein, B is a negative control.
FIG. 3: the antibacterial effect of the mixed lyase PlySs2, plySs2-PIGF fusion protein and collagen is shown in the figure. Wherein B is a negative control.
FIG. 4 is a schematic view of: bacteriostatic effect of different proteins and the mixture after being stored for 1 month at 4 ℃. Wherein, B is a negative control.
Detailed Description
The invention is further illustrated by the following specific examples in order to provide a better understanding of the invention, which are not to be construed as limiting the invention.
Example 1 preparation of lyase and lyase-collagen binding peptide fusion proteins
Firstly, taking lyase PlySs2 (WP _ 253220091.1) derived from a bacteriophage as an example, optimizing the amino acid sequence of the lyase PlySs2 according to an escherichia coli codon, integrating a DNA sequence between pET30a expression vectors NdeI and XhoI, and transforming the obtained vector into a BL21 (DE 3) expression strain to obtain a PlySs2 protein expression strain; and meanwhile, the amino acid sequence of PlySs2 is fused with the amino acid sequence of a collagen binding peptide PIGF2 mentioned in Mika ë lM. Martino research (namely the 123 th to 144 th amino acids of the PIGF-2, the specific sequence is RRPKGRGKRRREKQRPTDCHL), a DNA sequence is synchronously integrated between pET30a expression vectors NdeI and XhoI according to the codon optimization of escherichia coli, and the obtained vectors are transformed into BL21 (DE 3) expression strains and PlySs2-PIGF fusion protein expression strains.
Respectively transferring the above expression strains into LB culture medium for activation, culturing at 37 deg.C, and growing to OD 600 At around 0.8, IPTG was added to a final concentration of 0.5mM, and the mixture was incubated overnight at 220 rpm at 16 ℃. Centrifugally collecting the cultured overnight bacterial liquid at 7500 rpm, and resuspending the cells with 20 mM PB,150mM NaCl Buffer solution (Buffer for short) and pH 7.5; breaking cells with ultrasonic cell breaker, centrifuging cell breaking solution at 10000 rpm for more than 30 min, and collecting supernatant as crude enzyme solution. Then obtaining purified protein through His-Tag affinity chromatography, determining protein concentration through a photometric method, detecting the purity and the size of the protein by SDS-PAGE (polyacrylamide gel electrophoresis), wherein the result of figure 1 shows that each purified fusion protein has single band, the size is about 28-30KDa correctly and is consistent with the theoretical size, and the purity is evaluated>95％。
Example 2 determination of bacteriostatic Activity of proteins
The protein solution prepared in the above example 1 is used as a test object, the bacteriostatic activity of the lyase PlySs2 and PlySs2-PIGF fusion protein is detected, staphylococcus aureus GIM 1.481 is used as a bacteriostatic test strain, and the test is carried out by referring to a quantitative bacteriostatic experiment of WS/T650-2019.1.1 suspension, which is specifically as follows: the culture solution of staphylococcus aureus cultured overnight in LB medium at 37 ℃ is diluted 10000 times by 20 mM PB,150mM NaCl and pH 7.5 Buffer (Buffer for short), then added into PlySs2 and PlySs2-PIGF protein solutions respectively according to the addition of 2% (v/v), the concentration of the fusion protein of the proteins PlySs2 and PlySs2-PIGF is kept consistent and is 0.5mg/mL, the mixture is incubated at 37 ℃ for 1h, and then 100uL of the Buffer is taken for plating culture (LB medium). And diluting the staphylococcus aureus culture solution to a corresponding gradient by using a buffer solution for plating culture in a negative control mode, and observing and comparing the growth condition of staphylococcus aureus.
The results are shown in Table 1, and the number of Staphylococcus aureus on the negative control plate reaches 10 3 On the other hand, the numbers of Staphylococcus aureus on the sample plates treated with the lyase PlySs2 and the fusion protein PlySs2-PIGF are significantly reduced and almost all killed (FIG. 2). The result shows that the fusion of the collagen binding peptide and the lyase does not affect the bacteriostatic activity of the lyase on staphylococcus aureus, the bacteriostatic rates of the lyase and the staphylococcus aureus are all more than 99%, and the corresponding plate bacteriostatic effect graphs are shown in the figure.
TABLE 1 Staphylococcus aureus colony counts in the plates
Example 3 evaluation of the bacteriostatic effect and stability of fusion proteins of PlySs2 and PlySs2-PIGF with collagen
The protein samples prepared in example 1 above were mixed with collagen (recombinant human collagen derived from bernas) in equal volumes, each at a concentration of 0.5mg/mL. And (3) taking the mixture as a test object, and detecting the bacteriostatic activity of the mixed lyase PlySs2 and PlySs2-PIGF fusion protein and collagen, and evaluating the bacteriostatic activity after the mixture is placed for one month at 4 ℃ to characterize the stability of the mixture. Firstly, testing the antibacterial activity after mixing with collagen, taking staphylococcus aureus GIM 1.481 as an antibacterial test strain, and performing the test by referring to a WS/T650-2019.1.1 suspension quantitative antibacterial experiment, wherein the test method comprises the following specific steps: the culture solution of staphylococcus aureus cultured overnight in LB culture medium at 37 ℃ is diluted 10000 times by using 20 mM PB,150mM NaCl and pH 7.5 Buffer solution (Buffer for short), then the diluted culture solution is added into the compounded sample according to the addition of 2% (v/v), the compounded sample is placed at 37 ℃ for incubation for 1h, and then 100uL of the diluted culture solution is taken for plating culture (LB culture medium). And diluting the staphylococcus aureus culture solution to a corresponding gradient by using a buffer solution for negative control, and performing plate coating culture. The growth of Staphylococcus aureus was observed and compared.
The results are shown in Table 2, and the number of Staphylococcus aureus on the negative control plate reaches 10 4 On the other hand, the amount of staphylococcus aureus on the sample plate treated by the mixture formed by the lyase PlySs2, the fusion protein PlySs2-PIGF and the collagen is obviously reduced by more than 3 orders of magnitude. The results show that the lyase or the fusion protein and the collagen are mixed to achieve the killing effect on staphylococcus aureus, the bacteriostasis rate reaches over 99 percent, the collagen is endowed with new antibacterial activity, and the plate bacteriostasis effect diagram corresponding to the collagen mixture is shown in figure 3.
TABLE 2 Staphylococcus aureus colony counts in the plates
In the practical application process, the protein stability is an important characteristic of the bioactive substances and is also a key factor influencing the practical service life of the bioactive proteins, and the literature reports that the lyase PlySs2 protein can be stored at 4 ℃ for 15 days, and the protein is gradually inactivated after more than 15 days. In the invention, after the mixture is placed at 4 ℃ for one month, the mixture of PlySs2-PIGF fusion protein and collagen still keeps higher bacteriostatic activity, but the single lyase PlySs2 and collagen mixture is inactivated and is consistent with the activity cycle of the lyase PlySs2 reported in the literature, the specific bacteriostatic activity test method is the same as that of the negative control, the buffer solution is used for diluting the staphylococcus aureus culture solution to a corresponding gradient for plating culture, the other controls are respectively the lyase PlySs2 and the fusion protein PlySs2-PIGF solution, and the growth condition of the staphylococcus aureus is observed and compared.
The results of the experiment are shown in table 3.
TABLE 3 Staphylococcus aureus colony counts in the plates
As can be seen from the results in the table, the number of the negative control, plySs2 protein solution, fusion protein PlySs2-PIGF protein solution and the number of the staphylococcus aureus plate treated by the single lyase PlySs2 and collagen mixture reach 10 3 On the left and right, the number of staphylococcus aureus on a sample plate treated by the fusion protein PlySs2-PIGF collagen mixture is remarkably reduced, which shows that the protein per se is consistent with that reported by the literature, the storage period at 4 ℃ is short, the antibacterial activity on the staphylococcus aureus is lost after the sample plate is placed for one month, the fusion of the conjugated peptide PIGF does not change the stability of the protein per se, and the PlySs2 protein and the collagen are mixed into two independent individuals and do not improve the stability of the protein; after the fusion protein PlySs2-PIGF is mixed with collagen, the protein is combined with the collagen through PIGF binding peptide to form a complex, the stability of the protein is improved, the bacteriostatic result shows that the mixture still retains strong bacteriostatic activity after being placed for one month at 4 ℃, the bacteriostatic rate can reach more than 95 percent, and the corresponding plate bacteriostatic effect graph is shown in figure 4.
Claims (9)
1. A fusion peptide is characterized in that the fusion peptide is formed by fusing a lyase with a cell wall degradation effect and a collagen binding peptide; the lyase is bacteriophage lyase PlySs2 with NCBI accession number WP _253220091.1; the collagen binding peptide is collagen binding peptide PIGF2, and the amino acid sequence of the collagen binding peptide is RRPKGRGKRRREKQRPTDCHL.
2. An expression vector comprising the fusion peptide of claim 1.
3. A recombinant strain comprising an expression vector for the fusion peptide of claim 2.
4. The strain of claim 3, which is Escherichia coli.
5. A collagen complex having bacteriostatic activity, which is obtained by mixing the fusion peptide of claim 1 with collagen.
6. Use of the fusion peptide according to claim 1, or the collagen complex with bacteriostatic effect according to claim 5, for the preparation of a bacteriostatic product.
7. The use of claim 6, wherein the product is a pharmaceutical product, a cosmetic product, a toiletry product, a medical dressing, a medical tissue filler, a food package, an antimicrobial coating.
8. The use of claim 7, wherein the medicament is a pesticide.
9. The use of claim 7 or 8, wherein the inhibition of bacteria is inhibition of staphylococci.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211546201.2A CN115558031B (en) | 2022-12-05 | 2022-12-05 | Fusion peptide, collagen compound with antibacterial effect and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211546201.2A CN115558031B (en) | 2022-12-05 | 2022-12-05 | Fusion peptide, collagen compound with antibacterial effect and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN115558031A CN115558031A (en) | 2023-01-03 |
| CN115558031B true CN115558031B (en) | 2023-04-07 |
Family
ID=84770156
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211546201.2A Active CN115558031B (en) | 2022-12-05 | 2022-12-05 | Fusion peptide, collagen compound with antibacterial effect and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN115558031B (en) |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20050255043A1 (en) * | 2004-04-09 | 2005-11-17 | Hnatowich Donald J | Bacteriophage imaging of inflammation |
| CN102858985A (en) * | 2009-07-24 | 2013-01-02 | 西格马-奥尔德里奇有限责任公司 | Method for genome editing |
| EP4025698A1 (en) * | 2019-09-03 | 2022-07-13 | Sana Biotechnology, Inc. | Cd24-associated particles and related methods and uses thereof |
-
2022
- 2022-12-05 CN CN202211546201.2A patent/CN115558031B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN115558031A (en) | 2023-01-03 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| EP2283130B1 (en) | Dispersin B(TM), 5-fluorouracil, deoxyribonuclease I and proteinase K-based antibiofilm compositions and uses thereof | |
| US8821862B2 (en) | Soluble β-N-acetylglucosaminidase based antibiofilm compositions and uses thereof | |
| WO2008043175A1 (en) | SOLUBLE β-N-ACETYLGLUCOSAMINIDASE BASED ANTIBIOFILM COMPOSITIONS AND USES THEREOF | |
| JP2002544759A (en) | Compositions and methods for treating infections using cationic peptides alone or in combination with antibiotics | |
| AU2018231408B2 (en) | Phage therapy | |
| AU600886B2 (en) | Fibronectin-binding protein | |
| EP3852778A2 (en) | Bioactive peptides derived from snakes | |
| US10499655B2 (en) | Reagents and methods for inhibiting or disrupting biofilm | |
| CN101333258B (en) | Fusion polypeptide with antimicrobial and wound healing promoting function | |
| CN115558031B (en) | Fusion peptide, collagen compound with antibacterial effect and application thereof | |
| CN114159337B (en) | Facial antibacterial care solution containing recombinant human lysozyme | |
| KR20180031704A (en) | Combination therapy | |
| Shahzad et al. | Production of proteases by genetically improved Bacillus subtilis for enhanced skin penetration of antibacterial topical formulation | |
| JP2023545585A (en) | Combinations of novel bioactive peptides and their uses | |
| WO2005011587A2 (en) | Chalaropsis lysozyme protein and its method of use in anti-bacterial applications | |
| CN106707517B (en) | A kind of VR glasses | |
| Domínguez-Bello et al. | Resistance of rumen bacteria murein to bovine gastric lysozyme | |
| EP1328234A1 (en) | Methods for treating cystic fibrosis | |
| CN112625139B (en) | Protein and application thereof in promoting migration of skin fibroblasts, resisting bacteria and repairing wounds | |
| Fan et al. | Expression and purification of soluble porcine cystatin 11 in Pichia pastoris | |
| CN116262781B (en) | Antibacterial peptide descensin derivative, prokaryotic expression method and application thereof | |
| Alberdi Jr et al. | Antibacterial proteins from porcine polymorphonuclear neutrophils | |
| Salman et al. | The Role of Fructophilic Lactic Acid Bacteria in the Restoration of Infected Wounds and IL-17 Level in Rabbits | |
| Bharadwaz et al. | Isolation, purification, and characterization of staphylocoagulase, a blood coagulating protein from Staphylococcus sp. MBBJP S43 | |
| JPS62100298A (en) | Production of protein a |
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 |