CN113388624A - 一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法 - Google Patents
一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法 Download PDFInfo
- Publication number
- CN113388624A CN113388624A CN202011366261.7A CN202011366261A CN113388624A CN 113388624 A CN113388624 A CN 113388624A CN 202011366261 A CN202011366261 A CN 202011366261A CN 113388624 A CN113388624 A CN 113388624A
- Authority
- CN
- China
- Prior art keywords
- protein
- spaa
- identity
- erysipelas
- recombinant
- 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.)
- Pending
Links
- 108091007433 antigens Proteins 0.000 title claims abstract description 27
- 102000036639 antigens Human genes 0.000 title claims abstract description 27
- 208000024949 Swine Erysipelas Diseases 0.000 title claims abstract description 24
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 51
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 37
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 claims abstract description 14
- 201000000297 Erysipelas Diseases 0.000 claims abstract description 11
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 claims abstract description 11
- 239000000427 antigen Substances 0.000 claims abstract description 10
- 208000015181 infectious disease Diseases 0.000 claims abstract description 10
- 239000013604 expression vector Substances 0.000 claims abstract description 5
- 241000894006 Bacteria Species 0.000 claims description 26
- 229960005486 vaccine Drugs 0.000 claims description 15
- 239000013598 vector Substances 0.000 claims description 13
- 238000000034 method Methods 0.000 claims description 7
- 239000004094 surface-active agent Substances 0.000 claims description 4
- 239000000375 suspending agent Substances 0.000 claims description 4
- 241000282898 Sus scrofa Species 0.000 claims description 3
- 230000000890 antigenic effect Effects 0.000 claims description 3
- 239000003755 preservative agent Substances 0.000 claims description 3
- 239000003795 chemical substances by application Substances 0.000 claims description 2
- 238000001976 enzyme digestion Methods 0.000 claims description 2
- 230000000415 inactivating effect Effects 0.000 claims description 2
- 230000002335 preservative effect Effects 0.000 claims description 2
- 239000012646 vaccine adjuvant Substances 0.000 claims description 2
- 229940124931 vaccine adjuvant Drugs 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 241000588724 Escherichia coli Species 0.000 abstract description 5
- 108010076504 Protein Sorting Signals Proteins 0.000 abstract description 4
- 241000282887 Suidae Species 0.000 abstract description 4
- 108020004705 Codon Proteins 0.000 abstract description 3
- 238000005457 optimization Methods 0.000 abstract description 3
- 238000000746 purification Methods 0.000 abstract description 3
- 238000010367 cloning Methods 0.000 abstract description 2
- 230000001086 cytosolic effect Effects 0.000 abstract description 2
- 238000003745 diagnosis Methods 0.000 abstract description 2
- 244000144972 livestock Species 0.000 abstract description 2
- 238000011895 specific detection Methods 0.000 abstract description 2
- 241000186810 Erysipelothrix rhusiopathiae Species 0.000 description 21
- 239000002671 adjuvant Substances 0.000 description 21
- 239000007788 liquid Substances 0.000 description 18
- 239000013612 plasmid Substances 0.000 description 17
- 239000013641 positive control Substances 0.000 description 17
- 239000000243 solution Substances 0.000 description 17
- 239000006228 supernatant Substances 0.000 description 16
- RAXXELZNTBOGNW-UHFFFAOYSA-N imidazole Natural products C1=CNC=N1 RAXXELZNTBOGNW-UHFFFAOYSA-N 0.000 description 15
- 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 15
- 230000006698 induction Effects 0.000 description 13
- 230000001580 bacterial effect Effects 0.000 description 11
- 239000008188 pellet Substances 0.000 description 8
- 239000012880 LB liquid culture medium Substances 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 7
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 102000013566 Plasminogen Human genes 0.000 description 6
- 108010051456 Plasminogen Proteins 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 108020004414 DNA Proteins 0.000 description 5
- 150000001413 amino acids Chemical group 0.000 description 5
- 238000012258 culturing Methods 0.000 description 5
- 239000012154 double-distilled water Substances 0.000 description 5
- 239000000523 sample Substances 0.000 description 5
- QQPSCXKFDSORFT-IHRRRGAJSA-N Lys-Lys-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CCCCN QQPSCXKFDSORFT-IHRRRGAJSA-N 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 238000005119 centrifugation Methods 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 238000000108 ultra-filtration Methods 0.000 description 4
- 241000251468 Actinopterygii Species 0.000 description 3
- XFJKRRCWLTZIQA-XIRDDKMYSA-N Asn-Lys-Trp Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CC(=O)N)N XFJKRRCWLTZIQA-XIRDDKMYSA-N 0.000 description 3
- 102000018697 Membrane Proteins Human genes 0.000 description 3
- 108010052285 Membrane Proteins Proteins 0.000 description 3
- AJHCSUXXECOXOY-UHFFFAOYSA-N N-glycyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)CN)C(O)=O)=CNC2=C1 AJHCSUXXECOXOY-UHFFFAOYSA-N 0.000 description 3
- 239000003242 anti bacterial agent Substances 0.000 description 3
- 229940088710 antibiotic agent Drugs 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 238000007664 blowing Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 230000005847 immunogenicity Effects 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 3
- PRAKJMSDJKAYCZ-UHFFFAOYSA-N squalane Chemical compound CC(C)CCCC(C)CCCC(C)CCCCC(C)CCCC(C)CCCC(C)C PRAKJMSDJKAYCZ-UHFFFAOYSA-N 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 238000002525 ultrasonication Methods 0.000 description 3
- XQGIRPGAVLFKBJ-CIUDSAMLSA-N Ala-Asn-Lys Chemical compound N[C@@H](C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)O XQGIRPGAVLFKBJ-CIUDSAMLSA-N 0.000 description 2
- WNHNMKOFKCHKKD-BFHQHQDPSA-N Ala-Thr-Gly Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(O)=O WNHNMKOFKCHKKD-BFHQHQDPSA-N 0.000 description 2
- 229940046168 CpG oligodeoxynucleotide Drugs 0.000 description 2
- 108090000695 Cytokines Proteins 0.000 description 2
- 102000004127 Cytokines Human genes 0.000 description 2
- 108010088842 Fibrinolysin Proteins 0.000 description 2
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 2
- RVKIPWVMZANZLI-UHFFFAOYSA-N H-Lys-Trp-OH Natural products C1=CC=C2C(CC(NC(=O)C(N)CCCCN)C(O)=O)=CNC2=C1 RVKIPWVMZANZLI-UHFFFAOYSA-N 0.000 description 2
- 108010065920 Insulin Lispro Proteins 0.000 description 2
- 102100037850 Interferon gamma Human genes 0.000 description 2
- 108010074328 Interferon-gamma Proteins 0.000 description 2
- 241000880493 Leptailurus serval Species 0.000 description 2
- USLNHQZCDQJBOV-ZPFDUUQYSA-N Leu-Ile-Asn Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(O)=O USLNHQZCDQJBOV-ZPFDUUQYSA-N 0.000 description 2
- 241000699670 Mus sp. Species 0.000 description 2
- 108010087066 N2-tryptophyllysine Proteins 0.000 description 2
- 101710194807 Protective antigen Proteins 0.000 description 2
- UQFYNFTYDHUIMI-WHFBIAKZSA-N Ser-Gly-Ala Chemical compound OC(=O)[C@H](C)NC(=O)CNC(=O)[C@@H](N)CO UQFYNFTYDHUIMI-WHFBIAKZSA-N 0.000 description 2
- YSXYEJWDHBCTDJ-DVJZZOLTSA-N Thr-Gly-Trp Chemical compound C[C@H]([C@@H](C(=O)NCC(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)O)N)O YSXYEJWDHBCTDJ-DVJZZOLTSA-N 0.000 description 2
- GDPDVIBHJDFRFD-RNXOBYDBSA-N Trp-Tyr-Tyr Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O GDPDVIBHJDFRFD-RNXOBYDBSA-N 0.000 description 2
- HZZKQZDUIKVFDZ-AVGNSLFASA-N Tyr-Gln-Ser Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CO)C(=O)O)N)O HZZKQZDUIKVFDZ-AVGNSLFASA-N 0.000 description 2
- ILTXFANLDMJWPR-SIUGBPQLSA-N Tyr-Ile-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)N ILTXFANLDMJWPR-SIUGBPQLSA-N 0.000 description 2
- KSCVLGXNQXKUAR-JYJNAYRXSA-N Tyr-Leu-Glu Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O KSCVLGXNQXKUAR-JYJNAYRXSA-N 0.000 description 2
- HZDQUVQEVVYDDA-ACRUOGEOSA-N Tyr-Tyr-Leu Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(O)=O)NC(=O)[C@@H](N)CC=1C=CC(O)=CC=1)C1=CC=C(O)C=C1 HZDQUVQEVVYDDA-ACRUOGEOSA-N 0.000 description 2
- YQYFYUSYEDNLSD-YEPSODPASA-N Val-Thr-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(O)=O YQYFYUSYEDNLSD-YEPSODPASA-N 0.000 description 2
- KOSRFJWDECSPRO-UHFFFAOYSA-N alpha-L-glutamyl-L-glutamic acid Natural products OC(=O)CCC(N)C(=O)NC(CCC(O)=O)C(O)=O KOSRFJWDECSPRO-UHFFFAOYSA-N 0.000 description 2
- 230000004888 barrier function Effects 0.000 description 2
- 238000010009 beating Methods 0.000 description 2
- 230000010307 cell transformation Effects 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 231100000676 disease causative agent Toxicity 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000001035 drying Methods 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 239000000499 gel Substances 0.000 description 2
- 108010015792 glycyllysine Proteins 0.000 description 2
- 230000002480 immunoprotective effect Effects 0.000 description 2
- 210000003000 inclusion body Anatomy 0.000 description 2
- 230000001939 inductive effect Effects 0.000 description 2
- 108010044374 isoleucyl-tyrosine Proteins 0.000 description 2
- 108010057821 leucylproline Proteins 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 108010085203 methionylmethionine Proteins 0.000 description 2
- 239000000203 mixture Substances 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
- 239000003921 oil Substances 0.000 description 2
- 229940012957 plasmin Drugs 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000001742 protein purification Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 229940031626 subunit vaccine Drugs 0.000 description 2
- 108010061238 threonyl-glycine Proteins 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 108010073969 valyllysine Proteins 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- YYGNTYWPHWGJRM-UHFFFAOYSA-N (6E,10E,14E,18E)-2,6,10,15,19,23-hexamethyltetracosa-2,6,10,14,18,22-hexaene Chemical compound CC(C)=CCCC(C)=CCCC(C)=CCCC=C(C)CCC=C(C)CCC=C(C)C YYGNTYWPHWGJRM-UHFFFAOYSA-N 0.000 description 1
- ZORQXIQZAOLNGE-UHFFFAOYSA-N 1,1-difluorocyclohexane Chemical compound FC1(F)CCCCC1 ZORQXIQZAOLNGE-UHFFFAOYSA-N 0.000 description 1
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N Acrylic acid Chemical compound OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 1
- ZIWWTZWAKYBUOB-CIUDSAMLSA-N Ala-Asp-Leu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O ZIWWTZWAKYBUOB-CIUDSAMLSA-N 0.000 description 1
- OMMDTNGURYRDAC-NRPADANISA-N Ala-Glu-Val Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O OMMDTNGURYRDAC-NRPADANISA-N 0.000 description 1
- XUCHENWTTBFODJ-FXQIFTODSA-N Ala-Met-Ala Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](C)C(O)=O XUCHENWTTBFODJ-FXQIFTODSA-N 0.000 description 1
- NLOMBWNGESDVJU-GUBZILKMSA-N Ala-Met-Arg Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O NLOMBWNGESDVJU-GUBZILKMSA-N 0.000 description 1
- FVNAUOZKIPAYNA-BPNCWPANSA-N Ala-Met-Tyr Chemical compound CSCC[C@H](NC(=O)[C@H](C)N)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 FVNAUOZKIPAYNA-BPNCWPANSA-N 0.000 description 1
- WEZNQZHACPSMEF-QEJZJMRPSA-N Ala-Phe-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)C)CC1=CC=CC=C1 WEZNQZHACPSMEF-QEJZJMRPSA-N 0.000 description 1
- DFCIPNHFKOQAME-FXQIFTODSA-N Arg-Ala-Asn Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(N)=O)C(O)=O DFCIPNHFKOQAME-FXQIFTODSA-N 0.000 description 1
- DPXDVGDLWJYZBH-GUBZILKMSA-N Arg-Asn-Arg Chemical compound NC(N)=NCCC[C@H](N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCN=C(N)N)C(O)=O DPXDVGDLWJYZBH-GUBZILKMSA-N 0.000 description 1
- HKRXJBBCQBAGIM-FXQIFTODSA-N Arg-Asp-Ser Chemical compound C(C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CO)C(=O)O)N)CN=C(N)N HKRXJBBCQBAGIM-FXQIFTODSA-N 0.000 description 1
- UAOSDDXCTBIPCA-QXEWZRGKSA-N Arg-Ile-Gly Chemical compound CC[C@H](C)[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CCCN=C(N)N)N UAOSDDXCTBIPCA-QXEWZRGKSA-N 0.000 description 1
- JQHASVQBAKRJKD-GUBZILKMSA-N Arg-Ser-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CCCN=C(N)N)N JQHASVQBAKRJKD-GUBZILKMSA-N 0.000 description 1
- IOTKDTZEEBZNCM-UGYAYLCHSA-N Asn-Asn-Ile Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O IOTKDTZEEBZNCM-UGYAYLCHSA-N 0.000 description 1
- GNKVBRYFXYWXAB-WDSKDSINSA-N Asn-Glu-Gly Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O GNKVBRYFXYWXAB-WDSKDSINSA-N 0.000 description 1
- HDHZCEDPLTVHFZ-GUBZILKMSA-N Asn-Leu-Glu Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O HDHZCEDPLTVHFZ-GUBZILKMSA-N 0.000 description 1
- VOGCFWDZYYTEOY-DCAQKATOSA-N Asn-Lys-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CC(=O)N)N VOGCFWDZYYTEOY-DCAQKATOSA-N 0.000 description 1
- HPBNLFLSSQDFQW-WHFBIAKZSA-N Asn-Ser-Gly Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O HPBNLFLSSQDFQW-WHFBIAKZSA-N 0.000 description 1
- DATSKXOXPUAOLK-KKUMJFAQSA-N Asn-Tyr-Leu Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(O)=O DATSKXOXPUAOLK-KKUMJFAQSA-N 0.000 description 1
- HMQDRBKQMLRCCG-GMOBBJLQSA-N Asp-Arg-Ile Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O HMQDRBKQMLRCCG-GMOBBJLQSA-N 0.000 description 1
- XACXDSRQIXRMNS-OLHMAJIHSA-N Asp-Asn-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](CC(=O)O)N)O XACXDSRQIXRMNS-OLHMAJIHSA-N 0.000 description 1
- SNDBKTFJWVEVPO-WHFBIAKZSA-N Asp-Gly-Ser Chemical compound [H]N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CO)C(O)=O SNDBKTFJWVEVPO-WHFBIAKZSA-N 0.000 description 1
- YFSLJHLQOALGSY-ZPFDUUQYSA-N Asp-Ile-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CC(=O)O)N YFSLJHLQOALGSY-ZPFDUUQYSA-N 0.000 description 1
- GKWFMNNNYZHJHV-SRVKXCTJSA-N Asp-Lys-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CC(O)=O GKWFMNNNYZHJHV-SRVKXCTJSA-N 0.000 description 1
- ZXRQJQCXPSMNMR-XIRDDKMYSA-N Asp-Lys-Trp Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CC(=O)O)N ZXRQJQCXPSMNMR-XIRDDKMYSA-N 0.000 description 1
- MGSVBZIBCCKGCY-ZLUOBGJFSA-N Asp-Ser-Ser Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O MGSVBZIBCCKGCY-ZLUOBGJFSA-N 0.000 description 1
- NWAHPBGBDIFUFD-KKUMJFAQSA-N Asp-Tyr-Leu Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(O)=O NWAHPBGBDIFUFD-KKUMJFAQSA-N 0.000 description 1
- JGLWFWXGOINXEA-YDHLFZDLSA-N Asp-Val-Tyr Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 JGLWFWXGOINXEA-YDHLFZDLSA-N 0.000 description 1
- 206010060976 Bacillus infection Diseases 0.000 description 1
- 238000009007 Diagnostic Kit Methods 0.000 description 1
- 241000186811 Erysipelothrix Species 0.000 description 1
- 108700005480 Erysipelothrix rhusiopathiae SpaA Proteins 0.000 description 1
- 102000009123 Fibrin Human genes 0.000 description 1
- 108010073385 Fibrin Proteins 0.000 description 1
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 1
- PGPJSRSLQNXBDT-YUMQZZPRSA-N Gln-Arg-Gly Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O PGPJSRSLQNXBDT-YUMQZZPRSA-N 0.000 description 1
- WMOMPXKOKASNBK-PEFMBERDSA-N Gln-Asn-Ile Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O WMOMPXKOKASNBK-PEFMBERDSA-N 0.000 description 1
- PNENQZWRFMUZOM-DCAQKATOSA-N Gln-Glu-Leu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O PNENQZWRFMUZOM-DCAQKATOSA-N 0.000 description 1
- JHPFPROFOAJRFN-IHRRRGAJSA-N Gln-Glu-Tyr Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](CCC(=O)N)N)O JHPFPROFOAJRFN-IHRRRGAJSA-N 0.000 description 1
- GQZDDFRXSDGUNG-YVNDNENWSA-N Gln-Ile-Gln Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(N)=O)C(O)=O GQZDDFRXSDGUNG-YVNDNENWSA-N 0.000 description 1
- JRHPEMVLTRADLJ-AVGNSLFASA-N Gln-Lys-Lys Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)N)N JRHPEMVLTRADLJ-AVGNSLFASA-N 0.000 description 1
- QKWBEMCLYTYBNI-GVXVVHGQSA-N Gln-Lys-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CCC(N)=O QKWBEMCLYTYBNI-GVXVVHGQSA-N 0.000 description 1
- SXFPZRRVWSUYII-KBIXCLLPSA-N Gln-Ser-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(=O)N)N SXFPZRRVWSUYII-KBIXCLLPSA-N 0.000 description 1
- KPNWAJMEMRCLAL-GUBZILKMSA-N Gln-Ser-Lys Chemical compound C(CCN)C[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(=O)N)N KPNWAJMEMRCLAL-GUBZILKMSA-N 0.000 description 1
- KBKGRMNVKPSQIF-XDTLVQLUSA-N Glu-Ala-Tyr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O KBKGRMNVKPSQIF-XDTLVQLUSA-N 0.000 description 1
- LJLPOZGRPLORTF-CIUDSAMLSA-N Glu-Asn-Met Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCSC)C(O)=O LJLPOZGRPLORTF-CIUDSAMLSA-N 0.000 description 1
- RDDSZZJOKDVPAE-ACZMJKKPSA-N Glu-Asn-Ser Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CO)C(O)=O RDDSZZJOKDVPAE-ACZMJKKPSA-N 0.000 description 1
- HTTSBEBKVNEDFE-AUTRQRHGSA-N Glu-Gln-Val Chemical compound CC(C)[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCC(=O)O)N HTTSBEBKVNEDFE-AUTRQRHGSA-N 0.000 description 1
- IQACOVZVOMVILH-FXQIFTODSA-N Glu-Glu-Ser Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O IQACOVZVOMVILH-FXQIFTODSA-N 0.000 description 1
- ZWABFSSWTSAMQN-KBIXCLLPSA-N Glu-Ile-Ala Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(O)=O ZWABFSSWTSAMQN-KBIXCLLPSA-N 0.000 description 1
- CXRWMMRLEMVSEH-PEFMBERDSA-N Glu-Ile-Asn Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(O)=O CXRWMMRLEMVSEH-PEFMBERDSA-N 0.000 description 1
- PJBVXVBTTFZPHJ-GUBZILKMSA-N Glu-Leu-Asp Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)O)NC(=O)[C@H](CCC(=O)O)N PJBVXVBTTFZPHJ-GUBZILKMSA-N 0.000 description 1
- NWOUBJNMZDDGDT-AVGNSLFASA-N Glu-Leu-His Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CC1=CN=CN1 NWOUBJNMZDDGDT-AVGNSLFASA-N 0.000 description 1
- IVGJYOOGJLFKQE-AVGNSLFASA-N Glu-Leu-Lys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)O)N IVGJYOOGJLFKQE-AVGNSLFASA-N 0.000 description 1
- SWRVAQHFBRZVNX-GUBZILKMSA-N Glu-Lys-Asn Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(N)=O)C(O)=O SWRVAQHFBRZVNX-GUBZILKMSA-N 0.000 description 1
- DXVOKNVIKORTHQ-GUBZILKMSA-N Glu-Pro-Glu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(O)=O DXVOKNVIKORTHQ-GUBZILKMSA-N 0.000 description 1
- BFEZQZKEPRKKHV-SRVKXCTJSA-N Glu-Pro-Lys Chemical compound C1C[C@H](N(C1)C(=O)[C@H](CCC(=O)O)N)C(=O)N[C@@H](CCCCN)C(=O)O BFEZQZKEPRKKHV-SRVKXCTJSA-N 0.000 description 1
- ALMBZBOCGSVSAI-ACZMJKKPSA-N Glu-Ser-Asn Chemical compound C(CC(=O)O)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)N)C(=O)O)N ALMBZBOCGSVSAI-ACZMJKKPSA-N 0.000 description 1
- MRWYPDWDZSLWJM-ACZMJKKPSA-N Glu-Ser-Asp Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(O)=O)C(O)=O MRWYPDWDZSLWJM-ACZMJKKPSA-N 0.000 description 1
- IDEODOAVGCMUQV-GUBZILKMSA-N Glu-Ser-Leu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(O)=O IDEODOAVGCMUQV-GUBZILKMSA-N 0.000 description 1
- UCZXXMREFIETQW-AVGNSLFASA-N Glu-Tyr-Asn Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(N)=O)C(O)=O UCZXXMREFIETQW-AVGNSLFASA-N 0.000 description 1
- YMUFWNJHVPQNQD-ZKWXMUAHSA-N Gly-Ala-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)CN YMUFWNJHVPQNQD-ZKWXMUAHSA-N 0.000 description 1
- PHONXOACARQMPM-BQBZGAKWSA-N Gly-Ala-Met Chemical compound [H]NCC(=O)N[C@@H](C)C(=O)N[C@@H](CCSC)C(O)=O PHONXOACARQMPM-BQBZGAKWSA-N 0.000 description 1
- JXYMPBCYRKWJEE-BQBZGAKWSA-N Gly-Arg-Ala Chemical compound [H]NCC(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(O)=O JXYMPBCYRKWJEE-BQBZGAKWSA-N 0.000 description 1
- CCBIBMKQNXHNIN-ZETCQYMHSA-N Gly-Leu-Gly Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O CCBIBMKQNXHNIN-ZETCQYMHSA-N 0.000 description 1
- UHPAZODVFFYEEL-QWRGUYRKSA-N Gly-Leu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)CN UHPAZODVFFYEEL-QWRGUYRKSA-N 0.000 description 1
- DBJYVKDPGIFXFO-BQBZGAKWSA-N Gly-Met-Ala Chemical compound [H]NCC(=O)N[C@@H](CCSC)C(=O)N[C@@H](C)C(O)=O DBJYVKDPGIFXFO-BQBZGAKWSA-N 0.000 description 1
- WTUSRDZLLWGYAT-KCTSRDHCSA-N Gly-Trp-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)CN WTUSRDZLLWGYAT-KCTSRDHCSA-N 0.000 description 1
- LKJCZEPXHOIAIW-HOTGVXAUSA-N Gly-Trp-Lys Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)CN LKJCZEPXHOIAIW-HOTGVXAUSA-N 0.000 description 1
- FULZDMOZUZKGQU-ONGXEEELSA-N Gly-Val-His Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)CN FULZDMOZUZKGQU-ONGXEEELSA-N 0.000 description 1
- JBJNKUOMNZGQIM-PYJNHQTQSA-N His-Arg-Ile Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O JBJNKUOMNZGQIM-PYJNHQTQSA-N 0.000 description 1
- SDTPKSOWFXBACN-GUBZILKMSA-N His-Glu-Asp Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O SDTPKSOWFXBACN-GUBZILKMSA-N 0.000 description 1
- 108010093488 His-His-His-His-His-His Proteins 0.000 description 1
- GHAFKUCRIVBLDJ-IHRRRGAJSA-N His-His-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CC1=CN=CN1)NC(=O)[C@H](CC2=CN=CN2)N GHAFKUCRIVBLDJ-IHRRRGAJSA-N 0.000 description 1
- BZKDJRSZWLPJNI-SRVKXCTJSA-N His-His-Ser Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CO)C(O)=O BZKDJRSZWLPJNI-SRVKXCTJSA-N 0.000 description 1
- NKRJALPCDNXULF-BYULHYEWSA-N Ile-Asp-Gly Chemical compound [H]N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(O)=O NKRJALPCDNXULF-BYULHYEWSA-N 0.000 description 1
- BSWLQVGEVFYGIM-ZPFDUUQYSA-N Ile-Gln-Arg Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)N BSWLQVGEVFYGIM-ZPFDUUQYSA-N 0.000 description 1
- KMBPQYKVZBMRMH-PEFMBERDSA-N Ile-Gln-Asn Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O KMBPQYKVZBMRMH-PEFMBERDSA-N 0.000 description 1
- MTFVYKQRLXYAQN-LAEOZQHASA-N Ile-Glu-Gly Chemical compound [H]N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O MTFVYKQRLXYAQN-LAEOZQHASA-N 0.000 description 1
- PDTMWFVVNZYWTR-NHCYSSNCSA-N Ile-Gly-Lys Chemical compound CC[C@H](C)[C@H](N)C(=O)NCC(=O)N[C@@H](CCCCN)C(O)=O PDTMWFVVNZYWTR-NHCYSSNCSA-N 0.000 description 1
- IIWQTXMUALXGOV-PCBIJLKTSA-N Ile-Phe-Asp Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(=O)O)C(=O)O)N IIWQTXMUALXGOV-PCBIJLKTSA-N 0.000 description 1
- 102100026720 Interferon beta Human genes 0.000 description 1
- 108010047761 Interferon-alpha Proteins 0.000 description 1
- 102000006992 Interferon-alpha Human genes 0.000 description 1
- 108090000467 Interferon-beta Proteins 0.000 description 1
- 108010050904 Interferons Proteins 0.000 description 1
- 102000014150 Interferons Human genes 0.000 description 1
- 108010002352 Interleukin-1 Proteins 0.000 description 1
- 102000003814 Interleukin-10 Human genes 0.000 description 1
- 108090000174 Interleukin-10 Proteins 0.000 description 1
- 108010065805 Interleukin-12 Proteins 0.000 description 1
- 108010002350 Interleukin-2 Proteins 0.000 description 1
- 108090000978 Interleukin-4 Proteins 0.000 description 1
- 108010002616 Interleukin-5 Proteins 0.000 description 1
- 102000000743 Interleukin-5 Human genes 0.000 description 1
- 108010063738 Interleukins Proteins 0.000 description 1
- 102000015696 Interleukins Human genes 0.000 description 1
- PMGDADKJMCOXHX-UHFFFAOYSA-N L-Arginyl-L-glutamin-acetat Natural products NC(=N)NCCCC(N)C(=O)NC(CCC(N)=O)C(O)=O PMGDADKJMCOXHX-UHFFFAOYSA-N 0.000 description 1
- HGCNKOLVKRAVHD-UHFFFAOYSA-N L-Met-L-Phe Natural products CSCCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 HGCNKOLVKRAVHD-UHFFFAOYSA-N 0.000 description 1
- XBBKIIGCUMBKCO-JXUBOQSCSA-N Leu-Ala-Thr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O XBBKIIGCUMBKCO-JXUBOQSCSA-N 0.000 description 1
- MYGQXVYRZMKRDB-SRVKXCTJSA-N Leu-Asp-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN MYGQXVYRZMKRDB-SRVKXCTJSA-N 0.000 description 1
- VPKIQULSKFVCSM-SRVKXCTJSA-N Leu-Gln-Arg Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O VPKIQULSKFVCSM-SRVKXCTJSA-N 0.000 description 1
- DZQMXBALGUHGJT-GUBZILKMSA-N Leu-Glu-Ala Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(O)=O DZQMXBALGUHGJT-GUBZILKMSA-N 0.000 description 1
- YVKSMSDXKMSIRX-GUBZILKMSA-N Leu-Glu-Asn Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O YVKSMSDXKMSIRX-GUBZILKMSA-N 0.000 description 1
- HPBCTWSUJOGJSH-MNXVOIDGSA-N Leu-Glu-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O HPBCTWSUJOGJSH-MNXVOIDGSA-N 0.000 description 1
- QVFGXCVIXXBFHO-AVGNSLFASA-N Leu-Glu-Leu Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O QVFGXCVIXXBFHO-AVGNSLFASA-N 0.000 description 1
- HQUXQAMSWFIRET-AVGNSLFASA-N Leu-Glu-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN HQUXQAMSWFIRET-AVGNSLFASA-N 0.000 description 1
- XVZCXCTYGHPNEM-UHFFFAOYSA-N Leu-Leu-Pro Natural products CC(C)CC(N)C(=O)NC(CC(C)C)C(=O)N1CCCC1C(O)=O XVZCXCTYGHPNEM-UHFFFAOYSA-N 0.000 description 1
- LJBVRCDPWOJOEK-PPCPHDFISA-N Leu-Thr-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O LJBVRCDPWOJOEK-PPCPHDFISA-N 0.000 description 1
- XZNJZXJZBMBGGS-NHCYSSNCSA-N Leu-Val-Asn Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(N)=O)C(O)=O XZNJZXJZBMBGGS-NHCYSSNCSA-N 0.000 description 1
- YQFZRHYZLARWDY-IHRRRGAJSA-N Leu-Val-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCCN YQFZRHYZLARWDY-IHRRRGAJSA-N 0.000 description 1
- VKVDRTGWLVZJOM-DCAQKATOSA-N Leu-Val-Ser Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O VKVDRTGWLVZJOM-DCAQKATOSA-N 0.000 description 1
- 102000004083 Lymphotoxin-alpha Human genes 0.000 description 1
- 108090000542 Lymphotoxin-alpha Proteins 0.000 description 1
- NNCDAORZCMPZPX-GUBZILKMSA-N Lys-Gln-Ser Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CO)C(=O)O)N NNCDAORZCMPZPX-GUBZILKMSA-N 0.000 description 1
- IMAKMJCBYCSMHM-AVGNSLFASA-N Lys-Glu-Lys Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN IMAKMJCBYCSMHM-AVGNSLFASA-N 0.000 description 1
- ONPDTSFZAIWMDI-AVGNSLFASA-N Lys-Leu-Gln Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O ONPDTSFZAIWMDI-AVGNSLFASA-N 0.000 description 1
- LJADEBULDNKJNK-IHRRRGAJSA-N Lys-Leu-Val Chemical compound CC(C)C[C@H](NC(=O)[C@@H](N)CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O LJADEBULDNKJNK-IHRRRGAJSA-N 0.000 description 1
- ALGGDNMLQNFVIZ-SRVKXCTJSA-N Lys-Lys-Asp Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(=O)O)C(=O)O)N ALGGDNMLQNFVIZ-SRVKXCTJSA-N 0.000 description 1
- HVAUKHLDSDDROB-KKUMJFAQSA-N Lys-Lys-Leu Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O HVAUKHLDSDDROB-KKUMJFAQSA-N 0.000 description 1
- BXPHMHQHYHILBB-BZSNNMDCSA-N Lys-Lys-Tyr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O BXPHMHQHYHILBB-BZSNNMDCSA-N 0.000 description 1
- LECIJRIRMVOFMH-ULQDDVLXSA-N Lys-Pro-Phe Chemical compound NCCCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 LECIJRIRMVOFMH-ULQDDVLXSA-N 0.000 description 1
- YSPZCHGIWAQVKQ-AVGNSLFASA-N Lys-Pro-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)CCCCN YSPZCHGIWAQVKQ-AVGNSLFASA-N 0.000 description 1
- SBQDRNOLGSYHQA-YUMQZZPRSA-N Lys-Ser-Gly Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)NCC(O)=O SBQDRNOLGSYHQA-YUMQZZPRSA-N 0.000 description 1
- DLCAXBGXGOVUCD-PPCPHDFISA-N Lys-Thr-Ile Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O DLCAXBGXGOVUCD-PPCPHDFISA-N 0.000 description 1
- NROQVSYLPRLJIP-PMVMPFDFSA-N Lys-Trp-Tyr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O NROQVSYLPRLJIP-PMVMPFDFSA-N 0.000 description 1
- UGCIQUYEJIEHKX-GVXVVHGQSA-N Lys-Val-Glu Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O UGCIQUYEJIEHKX-GVXVVHGQSA-N 0.000 description 1
- NYTDJEZBAAFLLG-IHRRRGAJSA-N Lys-Val-Lys Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(O)=O NYTDJEZBAAFLLG-IHRRRGAJSA-N 0.000 description 1
- GILLQRYAWOMHED-DCAQKATOSA-N Lys-Val-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCCN GILLQRYAWOMHED-DCAQKATOSA-N 0.000 description 1
- 102000002274 Matrix Metalloproteinases Human genes 0.000 description 1
- 108010000684 Matrix Metalloproteinases Proteins 0.000 description 1
- HUKLXYYPZWPXCC-KZVJFYERSA-N Met-Ala-Thr Chemical compound CSCC[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O HUKLXYYPZWPXCC-KZVJFYERSA-N 0.000 description 1
- HLYIDXAXQIJYIG-CIUDSAMLSA-N Met-Gln-Asp Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O HLYIDXAXQIJYIG-CIUDSAMLSA-N 0.000 description 1
- VZBXCMCHIHEPBL-SRVKXCTJSA-N Met-Glu-Lys Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN VZBXCMCHIHEPBL-SRVKXCTJSA-N 0.000 description 1
- XKJUFUPCHARJKX-UWVGGRQHSA-N Met-Gly-His Chemical compound CSCC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CNC=N1 XKJUFUPCHARJKX-UWVGGRQHSA-N 0.000 description 1
- HLZORBMOISUNIV-DCAQKATOSA-N Met-Ser-Leu Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC(C)C HLZORBMOISUNIV-DCAQKATOSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- AUEJLPRZGVVDNU-UHFFFAOYSA-N N-L-tyrosyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CC1=CC=C(O)C=C1 AUEJLPRZGVVDNU-UHFFFAOYSA-N 0.000 description 1
- XMBSYZWANAQXEV-UHFFFAOYSA-N N-alpha-L-glutamyl-L-phenylalanine Natural products OC(=O)CCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 XMBSYZWANAQXEV-UHFFFAOYSA-N 0.000 description 1
- 108010079364 N-glycylalanine Proteins 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- BBDSZDHUCPSYAC-QEJZJMRPSA-N Phe-Ala-Leu Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(O)=O BBDSZDHUCPSYAC-QEJZJMRPSA-N 0.000 description 1
- UMKYAYXCMYYNHI-AVGNSLFASA-N Phe-Gln-Asn Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CC(=O)N)C(=O)O)N UMKYAYXCMYYNHI-AVGNSLFASA-N 0.000 description 1
- MGBRZXXGQBAULP-DRZSPHRISA-N Phe-Glu-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC1=CC=CC=C1 MGBRZXXGQBAULP-DRZSPHRISA-N 0.000 description 1
- HOYQLNNGMHXZDW-KKUMJFAQSA-N Phe-Glu-Arg Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O HOYQLNNGMHXZDW-KKUMJFAQSA-N 0.000 description 1
- GYEPCBNTTRORKW-PCBIJLKTSA-N Phe-Ile-Asp Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(O)=O GYEPCBNTTRORKW-PCBIJLKTSA-N 0.000 description 1
- RORUIHAWOLADSH-HJWJTTGWSA-N Phe-Ile-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CC1=CC=CC=C1 RORUIHAWOLADSH-HJWJTTGWSA-N 0.000 description 1
- XUSDDSLCRPUKLP-QXEWZRGKSA-N Pro-Asp-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H]1CCCN1 XUSDDSLCRPUKLP-QXEWZRGKSA-N 0.000 description 1
- LHALYDBUDCWMDY-CIUDSAMLSA-N Pro-Glu-Ala Chemical compound C[C@H](NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H]1CCCN1)C(O)=O LHALYDBUDCWMDY-CIUDSAMLSA-N 0.000 description 1
- AFXCXDQNRXTSBD-FJXKBIBVSA-N Pro-Gly-Thr Chemical compound [H]N1CCC[C@H]1C(=O)NCC(=O)N[C@@H]([C@@H](C)O)C(O)=O AFXCXDQNRXTSBD-FJXKBIBVSA-N 0.000 description 1
- YXHYJEPDKSYPSQ-AVGNSLFASA-N Pro-Leu-Arg Chemical compound NC(N)=NCCC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H]1CCCN1 YXHYJEPDKSYPSQ-AVGNSLFASA-N 0.000 description 1
- BRJGUPWVFXKBQI-XUXIUFHCSA-N Pro-Leu-Ile Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O BRJGUPWVFXKBQI-XUXIUFHCSA-N 0.000 description 1
- CPRLKHJUFAXVTD-ULQDDVLXSA-N Pro-Leu-Tyr Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O CPRLKHJUFAXVTD-ULQDDVLXSA-N 0.000 description 1
- ZLXKLMHAMDENIO-DCAQKATOSA-N Pro-Lys-Asp Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(O)=O)C(O)=O ZLXKLMHAMDENIO-DCAQKATOSA-N 0.000 description 1
- OOZJHTXCLJUODH-QXEWZRGKSA-N Pro-Val-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H]1CCCN1 OOZJHTXCLJUODH-QXEWZRGKSA-N 0.000 description 1
- KHRLUIPIMIQFGT-AVGNSLFASA-N Pro-Val-Leu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O KHRLUIPIMIQFGT-AVGNSLFASA-N 0.000 description 1
- 241000241413 Propolis Species 0.000 description 1
- 108010079005 RDV peptide Proteins 0.000 description 1
- 241000283984 Rodentia Species 0.000 description 1
- BRGQQXQKPUCUJQ-KBIXCLLPSA-N Ser-Glu-Ile Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O BRGQQXQKPUCUJQ-KBIXCLLPSA-N 0.000 description 1
- MUARUIBTKQJKFY-WHFBIAKZSA-N Ser-Gly-Asp Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O MUARUIBTKQJKFY-WHFBIAKZSA-N 0.000 description 1
- IXCHOHLPHNGFTJ-YUMQZZPRSA-N Ser-Gly-His Chemical compound C1=C(NC=N1)C[C@@H](C(=O)O)NC(=O)CNC(=O)[C@H](CO)N IXCHOHLPHNGFTJ-YUMQZZPRSA-N 0.000 description 1
- UAJAYRMZGNQILN-BQBZGAKWSA-N Ser-Gly-Met Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CCSC)C(O)=O UAJAYRMZGNQILN-BQBZGAKWSA-N 0.000 description 1
- DOSZISJPMCYEHT-NAKRPEOUSA-N Ser-Ile-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O DOSZISJPMCYEHT-NAKRPEOUSA-N 0.000 description 1
- UBRMZSHOOIVJPW-SRVKXCTJSA-N Ser-Leu-Lys Chemical compound OC[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(O)=O UBRMZSHOOIVJPW-SRVKXCTJSA-N 0.000 description 1
- GVMUJUPXFQFBBZ-GUBZILKMSA-N Ser-Lys-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(O)=O)C(O)=O GVMUJUPXFQFBBZ-GUBZILKMSA-N 0.000 description 1
- FPCGZYMRFFIYIH-CIUDSAMLSA-N Ser-Lys-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(O)=O FPCGZYMRFFIYIH-CIUDSAMLSA-N 0.000 description 1
- PMCMLDNPAZUYGI-DCAQKATOSA-N Ser-Lys-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O PMCMLDNPAZUYGI-DCAQKATOSA-N 0.000 description 1
- JUTGONBTALQWMK-NAKRPEOUSA-N Ser-Met-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CCSC)NC(=O)[C@H](CO)N JUTGONBTALQWMK-NAKRPEOUSA-N 0.000 description 1
- CKDXFSPMIDSMGV-GUBZILKMSA-N Ser-Pro-Val Chemical compound [H]N[C@@H](CO)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(O)=O CKDXFSPMIDSMGV-GUBZILKMSA-N 0.000 description 1
- VVKVHAOOUGNDPJ-SRVKXCTJSA-N Ser-Tyr-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CO)C(O)=O VVKVHAOOUGNDPJ-SRVKXCTJSA-N 0.000 description 1
- MFQMZDPAZRZAPV-NAKRPEOUSA-N Ser-Val-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CO)N MFQMZDPAZRZAPV-NAKRPEOUSA-N 0.000 description 1
- 229920002125 Sokalan® Polymers 0.000 description 1
- BHEOSNUKNHRBNM-UHFFFAOYSA-N Tetramethylsqualene Natural products CC(=C)C(C)CCC(=C)C(C)CCC(C)=CCCC=C(C)CCC(C)C(=C)CCC(C)C(C)=C BHEOSNUKNHRBNM-UHFFFAOYSA-N 0.000 description 1
- LMMDEZPNUTZJAY-GCJQMDKQSA-N Thr-Asp-Ala Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(O)=O LMMDEZPNUTZJAY-GCJQMDKQSA-N 0.000 description 1
- GKMYGVQDGVYCPC-IUKAMOBKSA-N Thr-Asp-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H]([C@@H](C)O)N GKMYGVQDGVYCPC-IUKAMOBKSA-N 0.000 description 1
- AHOLTQCAVBSUDP-PPCPHDFISA-N Thr-Ile-Lys Chemical compound CC[C@H](C)[C@H](NC(=O)[C@@H](N)[C@@H](C)O)C(=O)N[C@@H](CCCCN)C(O)=O AHOLTQCAVBSUDP-PPCPHDFISA-N 0.000 description 1
- 101000980463 Treponema pallidum (strain Nichols) Chaperonin GroEL Proteins 0.000 description 1
- UUIYFDAWNBSWPG-IHPCNDPISA-N Trp-Lys-Lys Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)O)N UUIYFDAWNBSWPG-IHPCNDPISA-N 0.000 description 1
- HSVPZJLMPLMPOX-BPNCWPANSA-N Tyr-Arg-Ala Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(O)=O HSVPZJLMPLMPOX-BPNCWPANSA-N 0.000 description 1
- IIJWXEUNETVJPV-IHRRRGAJSA-N Tyr-Arg-Ser Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CO)C(=O)O)N)O IIJWXEUNETVJPV-IHRRRGAJSA-N 0.000 description 1
- CKKFTIQYURNSEI-IHRRRGAJSA-N Tyr-Asn-Arg Chemical compound NC(N)=NCCC[C@@H](C(O)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CC1=CC=C(O)C=C1 CKKFTIQYURNSEI-IHRRRGAJSA-N 0.000 description 1
- HVHJYXDXRIWELT-RYUDHWBXSA-N Tyr-Glu-Gly Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O HVHJYXDXRIWELT-RYUDHWBXSA-N 0.000 description 1
- UNUZEBFXGWVAOP-DZKIICNBSA-N Tyr-Glu-Val Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O UNUZEBFXGWVAOP-DZKIICNBSA-N 0.000 description 1
- 102000003990 Urokinase-type plasminogen activator Human genes 0.000 description 1
- 108090000435 Urokinase-type plasminogen activator Proteins 0.000 description 1
- PAPWZOJOLKZEFR-AVGNSLFASA-N Val-Arg-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CCCCN)C(=O)O)N PAPWZOJOLKZEFR-AVGNSLFASA-N 0.000 description 1
- ZMDCGGKHRKNWKD-LAEOZQHASA-N Val-Asn-Glu Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N ZMDCGGKHRKNWKD-LAEOZQHASA-N 0.000 description 1
- VLOYGOZDPGYWFO-LAEOZQHASA-N Val-Asp-Glu Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O VLOYGOZDPGYWFO-LAEOZQHASA-N 0.000 description 1
- QHDXUYOYTPWCSK-RCOVLWMOSA-N Val-Asp-Gly Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)NCC(=O)O)N QHDXUYOYTPWCSK-RCOVLWMOSA-N 0.000 description 1
- TZVUSFMQWPWHON-NHCYSSNCSA-N Val-Asp-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](C(C)C)N TZVUSFMQWPWHON-NHCYSSNCSA-N 0.000 description 1
- JAKHAONCJJZVHT-DCAQKATOSA-N Val-Lys-Ser Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)O)N JAKHAONCJJZVHT-DCAQKATOSA-N 0.000 description 1
- KSFXWENSJABBFI-ZKWXMUAHSA-N Val-Ser-Asn Chemical compound [H]N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O KSFXWENSJABBFI-ZKWXMUAHSA-N 0.000 description 1
- VIKZGAUAKQZDOF-NRPADANISA-N Val-Ser-Glu Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCC(O)=O VIKZGAUAKQZDOF-NRPADANISA-N 0.000 description 1
- RTJPAGFXOWEBAI-SRVKXCTJSA-N Val-Val-Arg Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCN=C(N)N RTJPAGFXOWEBAI-SRVKXCTJSA-N 0.000 description 1
- NLNCNKIVJPEFBC-DLOVCJGASA-N Val-Val-Glu Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCC(O)=O NLNCNKIVJPEFBC-DLOVCJGASA-N 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 238000001261 affinity purification Methods 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 108010044940 alanylglutamine Proteins 0.000 description 1
- 108010087924 alanylproline Proteins 0.000 description 1
- 108010070783 alanyltyrosine Proteins 0.000 description 1
- AZDRQVAHHNSJOQ-UHFFFAOYSA-N alumane Chemical class [AlH3] AZDRQVAHHNSJOQ-UHFFFAOYSA-N 0.000 description 1
- CEGOLXSVJUTHNZ-UHFFFAOYSA-K aluminium tristearate Chemical compound [Al+3].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O CEGOLXSVJUTHNZ-UHFFFAOYSA-K 0.000 description 1
- 229940024546 aluminum hydroxide gel Drugs 0.000 description 1
- 229940063655 aluminum stearate Drugs 0.000 description 1
- SMYKVLBUSSNXMV-UHFFFAOYSA-K aluminum;trihydroxide;hydrate Chemical compound O.[OH-].[OH-].[OH-].[Al+3] SMYKVLBUSSNXMV-UHFFFAOYSA-K 0.000 description 1
- 239000010775 animal oil Substances 0.000 description 1
- 108010008355 arginyl-glutamine Proteins 0.000 description 1
- 108010068380 arginylarginine Proteins 0.000 description 1
- 108010038633 aspartylglutamate Proteins 0.000 description 1
- 108010068265 aspartyltyrosine Proteins 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- VEZXCJBBBCKRPI-UHFFFAOYSA-N beta-propiolactone Chemical compound O=C1CCO1 VEZXCJBBBCKRPI-UHFFFAOYSA-N 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 229960001631 carbomer Drugs 0.000 description 1
- 230000007969 cellular immunity Effects 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000003021 clonogenic effect Effects 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 108010054813 diprotin B Proteins 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 229950003499 fibrin Drugs 0.000 description 1
- 239000003527 fibrinolytic agent Substances 0.000 description 1
- 230000003480 fibrinolytic effect Effects 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 108010055341 glutamyl-glutamic acid Proteins 0.000 description 1
- VPZXBVLAVMBEQI-UHFFFAOYSA-N glycyl-DL-alpha-alanine Natural products OC(=O)C(C)NC(=O)CN VPZXBVLAVMBEQI-UHFFFAOYSA-N 0.000 description 1
- XBGGUPMXALFZOT-UHFFFAOYSA-N glycyl-L-tyrosine hemihydrate Natural products NCC(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 XBGGUPMXALFZOT-UHFFFAOYSA-N 0.000 description 1
- 108010050848 glycylleucine Proteins 0.000 description 1
- 108010087823 glycyltyrosine Proteins 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 108010040030 histidinoalanine Proteins 0.000 description 1
- 108010028295 histidylhistidine Proteins 0.000 description 1
- 108010085325 histidylproline Proteins 0.000 description 1
- 244000052637 human pathogen Species 0.000 description 1
- 230000004727 humoral immunity Effects 0.000 description 1
- 239000012642 immune effector Substances 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 229940121354 immunomodulator Drugs 0.000 description 1
- 230000003308 immunostimulating effect Effects 0.000 description 1
- 230000000937 inactivator Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 229940047124 interferons Drugs 0.000 description 1
- 229940047122 interleukins Drugs 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 231100000518 lethal Toxicity 0.000 description 1
- 230000001665 lethal effect Effects 0.000 description 1
- 108010034529 leucyl-lysine Proteins 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 239000012160 loading buffer Substances 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 108010009298 lysylglutamic acid Proteins 0.000 description 1
- 108010064235 lysylglycine Proteins 0.000 description 1
- 108010054155 lysyllysine Proteins 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 108010068488 methionylphenylalanine Proteins 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 210000003097 mucus Anatomy 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- JXTPJDDICSTXJX-UHFFFAOYSA-N n-Triacontane Natural products CCCCCCCCCCCCCCCCCCCCCCCCCCCCCC JXTPJDDICSTXJX-UHFFFAOYSA-N 0.000 description 1
- OHDXDNUPVVYWOV-UHFFFAOYSA-N n-methyl-1-(2-naphthalen-1-ylsulfanylphenyl)methanamine Chemical compound CNCC1=CC=CC=C1SC1=CC=CC2=CC=CC=C12 OHDXDNUPVVYWOV-UHFFFAOYSA-N 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 229940060184 oil ingredients Drugs 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 108010073101 phenylalanylleucine Proteins 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 238000002264 polyacrylamide gel electrophoresis Methods 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000001376 precipitating effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 229960000380 propiolactone Drugs 0.000 description 1
- 229940069949 propolis Drugs 0.000 description 1
- 230000004224 protection Effects 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 239000012474 protein marker Substances 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 208000017520 skin disease Diseases 0.000 description 1
- 239000001593 sorbitan monooleate Substances 0.000 description 1
- 229940035049 sorbitan monooleate Drugs 0.000 description 1
- 235000011069 sorbitan monooleate Nutrition 0.000 description 1
- 229940032094 squalane Drugs 0.000 description 1
- 229940031439 squalene Drugs 0.000 description 1
- TUHBEKDERLKLEC-UHFFFAOYSA-N squalene Natural products CC(=CCCC(=CCCC(=CCCC=C(/C)CCC=C(/C)CC=C(C)C)C)C)C TUHBEKDERLKLEC-UHFFFAOYSA-N 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000013595 supernatant sample Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 238000009210 therapy by ultrasound Methods 0.000 description 1
- RTKIYNMVFMVABJ-UHFFFAOYSA-L thimerosal Chemical compound [Na+].CC[Hg]SC1=CC=CC=C1C([O-])=O RTKIYNMVFMVABJ-UHFFFAOYSA-L 0.000 description 1
- 229940033663 thimerosal Drugs 0.000 description 1
- 230000000699 topical effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 108010044292 tryptophyltyrosine Proteins 0.000 description 1
- 108010020532 tyrosyl-proline Proteins 0.000 description 1
- 108010003137 tyrosyltyrosine Proteins 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 229960005356 urokinase Drugs 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 230000007923 virulence factor Effects 0.000 description 1
- 239000000304 virulence factor Substances 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
- 206010048282 zoonosis Diseases 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
- C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/02—Bacterial antigens
- A61K39/0241—Mollicutes, e.g. Mycoplasma, Erysipelothrix
-
- 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
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/55—Medicinal preparations containing antigens or antibodies characterised by the host/recipient, e.g. newborn with maternal antibodies
- A61K2039/552—Veterinary vaccine
-
- 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
- C12N2800/22—Vectors comprising a coding region that has been codon optimised for expression in a respective host
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Zoology (AREA)
- Animal Behavior & Ethology (AREA)
- Wood Science & Technology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Microbiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- Immunology (AREA)
- Epidemiology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Physics & Mathematics (AREA)
- Oncology (AREA)
- Communicable Diseases (AREA)
- Gastroenterology & Hepatology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Plant Pathology (AREA)
- Mycology (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Peptides Or Proteins (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
本发明涉及一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法,所述猪丹毒SpaA抗原蛋白的编码基因序列如SEQ ID NO.1所示;所述制备方法包括如下步骤:构建含有上述重组蛋白基因的表达载体;重组蛋白的表达。本发明通过分析SpaA蛋白序列,根绝其序列特征进行了剔除和优化;将信号肽等序列删除,通过兼并密码子优化获得克隆序列;并利用大肠杆菌的胞质空间进行克隆和蛋白表达,最后进行纯化获得重组SpaA蛋白;抗原包被后进行畜牧猪丹毒感染的特异性检测,为后续诊断和治疗提供了基础。
Description
技术领域
本发明涉及疫苗技术领域,尤其是涉及一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法。
背景技术
猪丹毒杆菌(Erysipelothrix rhusiopathiae)是一种细长的,多形性,无孢子的革兰氏阳性杆菌,直径为0.2-0.4pm,长度为0.8-2.5μm。菌体以单链、短链、双链呈“V”形或随机组合排列,有时也会以丝状体或长链排列。猪丹毒杆菌的最佳生长温度在30-37℃,但它的生长温度范围可到5-44℃。其最佳的生长pH值在7.2-7.6,但生长pH值范围为6.8-8.2,是一种厌氧菌。于1878年由koch首次分离出来。到1886年,Loeffler检测认定其为猪丹毒疾病的病原体。1909年,Rosenbach又从人类中的局部皮肤病患者身上分离得到这种菌,从而确认了它也是一种人类病原体,因此,猪丹毒杆菌是一类人畜共患类疾病——猪丹毒的病原体。猪丹毒杆菌的主要宿主一般是家猪,但也会感染其他多种鸟类和啮齿类动物,鱼类的感染虽然不会出现明确的已知病症,但是猪丹毒杆菌能在鱼类的体表粘液中长期生存,也就是说鱼类能够长期携带猪丹毒杆菌,这大大增加了人类感染猪丹毒杆菌的风险。
近几年的研究发现,猪丹毒杆菌中存在几种相对分子质量为64、66、43kDa的表面蛋白具有免疫保护作用,Makino研究组克隆表达出大小为64kDa的表面蛋白,将其命名为表面保护性抗原A,即猪丹毒SpaA蛋白(surface protein A)。多个研究小组对SpaA蛋白进行免疫功能的研究后认定SpaA蛋白具有良好的免疫保护效用。随后Ho To等又研究发现SpaA蛋白是11种主要血清型的猪丹毒杆菌的共有抗原,这个研究表明了SpaA蛋白广泛存在于多种猪丹毒杆菌菌株中。Kitajima等从血清型2猪丹毒杆菌Kyoto株种提取抗原SpaA蛋白,然后用血清型1a的强毒株和血清型2的强毒株进行肌肉攻击,结果表明从血清型2的菌株种提取的SpaA抗原能够完全保护猪受到同源强毒株或异源强毒株的致死性感染,并且其免疫保护能力强于传统的弱毒活菌疫苗,这个实验也一定程度证明了SpaA蛋白具有对多种血清型的猪丹毒杆菌产生免疫原性的潜力,是制备猪丹毒杆菌亚单位疫苗的候选抗原。编码的SpaA由N端的免疫保护性区域和C端的细胞结合区域组成。重组SpaA可诱导产生较高水平的特异性抗体IgG,使Th1、Th2细胞因子 (TNF-β、IFN-γ、IL-5、IL-10)含量显著提升。因此,SpaA能共同激发机体的体液免疫和细胞免疫。小鼠保护性试验及病理组织学观察研究表明,在存在尿激酶的情况下,猪丹毒杆菌可以结合宿主纤溶酶原并表现出纤溶酶活性,将纤溶酶原系统用于跨组织屏障的迁移或感染期间的营养需求。因此,像其他病原菌一样,猪丹毒杆菌可在以下四个方面利用纤溶酶原/纤溶酶系统作为毒力因子:靶向宿主纤溶系统和降解纤维蛋白凝块,产生生物活性裂解片段以影响信号传导途径,激活基质金属蛋白酶以协助降解组织屏障,并破坏免疫效应分子以逃避免疫。因此,纤维蛋白溶酶原的利用可能是在猪丹毒杆菌感染过程中的重要机制。而SpaA可以特异性结合宿主纤溶酶原,因此抗SpaA血清可显著降低猪丹毒杆菌结合纤溶酶原的活性。其他一些研究又表明SpaA蛋白参与了猪丹毒杆菌宿主细胞的粘附以及对补体介导的杀伤的抑制。综上所述,SpaA蛋白能有效干预猪丹毒杆菌对宿主的感染,具有良好的免疫保护作用,有成为猪丹毒的新型亚单位疫苗的巨大潜力,基于目前猪丹毒杆菌仍旧无法得到良好有效的控制的情况,这让SpaA蛋白成为了目前猪丹毒新型疫苗的主要研究方向。目前针对猪丹毒检测和治理仍然没有有效的疫苗和诊断试剂盒,主要是由于猪丹毒SpaA序列比较长,因而比较难以获得有效抗原序列。
CN201910442887.2公开了一种猪丹毒SpaA蛋白及其在制备疫苗中的应用。CN200580013595.3公开了一种防止猪丹毒杆菌感染的猪丹毒杆菌表面保护性抗原SpaA蛋白或其中SpaA蛋白被除去一部分后得到的缩短型SpaA蛋白的变体及其制备方法。在SpaA蛋白或SpaA蛋白的氨基酸序列的特定位点上引入氨基酸取代得到具有免疫原性,以包涵体形式在大肠杆菌中表达的SpaA蛋白或SpaA蛋白变体。由于本发明SpaA蛋白或SpaA蛋白变体以包涵体形式在大肠杆菌中表达,可容易地回收和纯化。
发明内容
为解决上述技术问题,本发明的上述目的通过以下技术方案来实现:
一种猪丹毒SpaA抗原蛋白,其特征在于,其编码基因序列与SEQ ID NO.1具有大于90%的同一性,或大于92%的同一性,或大于95%的同一性,或大于96%的同一性,或大于97%的同一性,或大于98%的同一性,或大于99%的同一性;优选如SEQ ID NO.1所示。
一种猪丹毒SpaA抗原蛋白,其特征在于,其氨基酸序列与SEQ ID NO.2具有大于90%的同一性,或大于92%的同一性,或大于95%的同一性,或大于96%的同一性,或大于97%的同一性,或大于98%的同一性,或大于99%的同一性;优选如SEQ ID NO.2所示。
一种重组载体,其特征在于,含有上述猪丹毒SpaA抗原蛋白的编码基因;优选是pET-16b-ErSpaA。
一种猪丹毒SpaA基因工程疫苗,其特征在于,所述疫苗包含上述猪丹毒SpaA抗原蛋白;或包含含有上述猪丹毒SpaA抗原蛋白的编码基因的重组载体。
其中一些实施例,所述疫苗还包含疫苗佐剂,如兽医学可接受的水性佐剂或油性佐剂。优选的,水性佐剂包括但不限于铝盐系列佐剂、Montanide IMS系列佐剂或MontanideGEL系列佐剂、蜂胶、免疫刺激复合物、细胞因子类佐剂、核酸及其衍生物类佐剂、卵磷脂类佐剂。所述Montanide IMS系列佐剂包括1313VG、251C VG、2215VG;所述Montanide GEL系列佐剂为GEL 01PR或Montanide PET GEL A;所述水包油性系列佐剂包括MF59、MontanideISA15A VG等;所述细胞因子类佐剂包括白介素(IL-1、IL-2、IL-4、IL-12),干扰素(IFN-γ、IFN-α、IFN-β)等;所述核酸及其衍生物类佐剂包括免疫刺激序列DNA(CpG DNA)或CpG寡聚脱氧核苷酸等。进一步优选的,水性佐剂优选方式涉及为IMS1313N VG、IMS2215VG、Gel01、卡波姆、氢氧化铝胶的一种或几种的组合物。其中一些实施例,所述疫苗包含弗式完全佐剂和/或弗式不完全佐剂。
其中一些实施例,所述佐剂的浓度范围是从5~50%V/V,优选20~30%V/V,进一步优选25% V/V。
其中一些实施例,所述油性佐剂包括但不限于白油、角鲨烷或角鲨烯、德雷克油(Drakeoil),以及其他动物油、植物油或矿物油。上述油性佐剂既可以是天然来源,也可以是经过人工合成获得的。
其中一些实施例,所述疫苗还包括助悬剂、表面活性剂、抗原灭活剂或防腐剂。所述助悬剂可包括,例如,硬脂酸铝,以及所属技术领域可用的其他助悬剂。所述表面活性剂可包括,例如,脱水山梨醇单油酸酯(TWEEN系列),司盘(SPAN系列),以及所属技术领域可用的其他表面活性剂。所述抗原灭活剂包括但不限于,例如福尔马林、β-丙内酯等等。所述防腐剂包括,例如硫柳汞。上述物质的使用方法及用量均为本领域技术人员所熟知。
本发明还提供一种猪丹毒SpaA抗原蛋白优化克隆的制备方法,其特征在于,包括如下步骤:
构建含有上述重组蛋白基因的表达载体;
重组蛋白的表达。
其中一些实施例,上述制备方法还包括对表达系统表达重组蛋白的纯化的步骤。
其中一些实施例,上述制备方法还包括在构建表达载体前对目的基因的获取步骤。
其中一些实施例,上述制备方法的具体步骤为:
将获取的目的基因酶切后连接入pET-16b载体,转入克隆菌,获得重组载体pET-16b-ErSpaA;
上述重组载体分别转入表达菌中,诱导表达目的蛋白。
其中一些实施例,克隆菌优选甘油菌,表达菌优选Rossetta gami B或BL21 star。
本发明还提供了上述猪丹毒SpaA抗原蛋白、上述重组载体、上述疫苗在制备抗猪丹毒杆菌感染的疫苗中的应用。
本发明具有以下有益的效果:本发明通过分析SpaA蛋白序列,根绝其序列特征进行了剔除和优化;将信号肽等序列删除,通过兼并密码子优化获得克隆序列;并利用大肠杆菌的胞质空间进行克隆和蛋白表达,最后进行纯化获得重组SpaA蛋白;抗原包被后进行畜牧猪丹毒感染的特异性检测,为后续诊断和治疗提供了基础。
附图说明
图1是pET-16b-ErSpaA重组质粒鉴定结果(M:DL 15000 DNA Maker;1:pET-16b-ErSpaA质粒;2:pET-16b-ErSpaA质粒;3:pET-16b-ErSpaA质粒),其中浓度测量结果分别为:
1号:浓度44.1 ng/μL 260/280=1.99;
2号:浓度49.4 ng/μL 260/280=1.92;
3号:浓度42.5 ng/μL 260/280=1.95。
图2是pET-16b-ErSpaA重组质粒构建示意图。
图3是猪丹毒杆菌SpaA基因的PCR鉴定结果(M:DL2000 DNA MAKER;1:pET-16b-ErSpaA-Rossetta gami B(DE3)重组菌菌液PCR;2:pET-16b-ErSpaA质粒PCR(阳性对照))。
图4是蛋白样制备流程图。
图5是重组菌表达鉴定结果,M:蛋白Maker C600525;1:BL21 star未诱导表达;2:BL21 star诱导后原样;3:BL21 star诱导后沉淀;4:BL21 star诱导后上清;5:Rossettagami B未诱导表达;6:Rossetta gami B诱导后原样;7:Rossetta gami B诱导后沉淀;8:Rossetta gami B诱导后上清。
图6是蛋白质纯化结果。
具体实施方式
下面通过具体实施方式来进一步说明本发明的技术方案。本领域技术人员应施例仅仅是帮助理解本发明,不应视为对本发明的具体限制。
本发明所述技术方案中,如未特别说明,均为本领域的常规技术;所述试剂或材料,如未特别说明,均购自商业渠道。
实验材料:pET16b-ErSpaA质粒及TOP10重组菌(南京金斯瑞合成)、引物SpaA1(aaggatccGACAGCACCGATATCAGC)/SpaA2(agctcgagTTATTTCAGGCTGCCGTC)(上海生工)、BL21star菌株(invitrogen)、Rossetta gami B菌株(上海生工)、非感受态细胞转化试剂盒(上海生工)、2×PCR Master(上海生工)、Amp抗生素(石药集团)、Plasmid Miniprep Kit(Biomiga)、PAGE凝胶快速制备试剂盒(雅酶生物)、Ni2+ NTA beads亲和层析填料(常州天地人和)、30 kD超滤管(millipore)、咪唑、1 M Tis-HCl(pH8.0)、NaCl、蛋白胨、酵母膏、IPTG。
实验仪器:摇床、超净台、移液枪、高速离心机、超声破碎仪、电泳仪等。
实施例1 ErSpaA基因合成及重组质粒构建
ErSpaA基因编码606个氨基酸,其中包括29个氨基酸的信号肽。将编码信号肽的核苷酸序列去除,得到长为1734 bp的编码成熟ErSpaA的全序列基因,序列如SEQ ID NO.1所示,完成大肠杆菌密码子优化并合成,构建得到重组质粒pET-16b-ErSpaA(见图1和图2)。
实施例2 pET-16b-ErSpaA转化BL21 star、Rossetta gami B
a. 接种甘油菌
取构建好的甘油菌pET-16b-ErSpaA Top 10,分别接种100μL菌液到两管5 mL LB液体培养基,同时加50 mg/mL Amp 5μL(终浓度为50 μg/mL)。一管做pET-16b-ErSpaA质粒提取,用Plasmid Miniprep Kit试剂盒按操作步骤提取pET-16b-ErSpaA质粒,另一管保存。
b. 非感受态细胞转化
分别接种500μL BL21 star、Rossetta gami B菌种至5mL LB液体培养基中,在37℃,220 rpm的条件下摇床培养2h;随后于超净台取1mL菌液,5000 rpm离心5 min,小心丢弃上清;短暂离心将管壁液体甩至管底,用移液器将残余的液体培养基吸尽;加入50μL冰上预冷的BT Buffer K,用移液枪将菌体沉淀轻轻吹打混匀;加入5μL pET-16b-ErSpaA质粒,轻轻混匀;将上述离心管置于冰上放置5 min;然后加入1 mL预热至37℃的无抗生素的LB液体培养基,混匀后于37℃条件下摇床培养45- 60 min;将上述1 mL菌液接种至4mL LB液体培养基中,并加入50 mg/mL的Amp 5μL,37℃,220 rpm条件下摇床培养,待菌长出。
鉴定转化成功后,三区划线分离单菌落。
c. 菌液PCR鉴定
首先制备模板:超净台取50μL 上述菌液;5000 rmp离心5 min后,弃上清,用移液枪将残余培养基吸干,然后加200μL ddH2O重悬;再5000 rmp离心5 min,弃上清,用枪吸干,加200μL ddH2O重悬;然后在100℃中煮10 min 煮样;5000 rmp离心,取上清到新的EP管中,即为模板。
然后溶解引物:将引物粉末配平离心;SpaA1加770μL ddH2O,SpaA2加830μL ddH2O并吹打溶解;涡旋(点动)5次左右;将管壁上的液体甩下去即可使用。
进行PCR:
2×PCR Master 20 μL
SpaA1 2 μL
SpaA2 2 μL
ddH2O 14 μL
最后各加入模板1μL,混匀。PCR检测中实验组为上述制备的模板,阳性对照为质粒pET-16b-ErSpaA。
PCR鉴定结果如下(见图3):以所提取的菌株DNA为模板,进行重组SpaA基因的PCR扩增。作为阳性对照的pET-16b-ErSpaA质粒PCR得到了预期的1734bp的目的条带。pET-16b-ErSpaA/Rossetta gami B重组菌菌液的PCR结果与阳性对照相对比,同样得到了预期的1734bp的目的条带。证明了实验中pET-16b-ErSpaA质粒成功转化非感受态细胞。
PCR反应条件为:95℃ 5min;然后95℃ 15s;58℃ 15s;72℃ 2min进行30个循环;72℃ 10min。PCR产物用10g/L琼脂糖凝胶电泳,回收后对纯化的PCR产物进行测定。
实施例3 IPTG诱导表达重组菌
经鉴定成功的重组菌,划线筛选单菌落并保存。分别将pET-16b-ErSpaA/BL21star、pET-16b-ErSpaA/Rossetta gami B重组菌接种100μL到两管5 mL的LB液体培养基中,同时各加入50 mg/mL的Amp 5μL,一管加IPTG诱导表达,一管不诱导作为对照。
37℃,220 rpm摇床培养约6 h后,其中一管加5μL 1M IPTG进行诱导,继续37℃,220 rpm摇床培养4 h。IPTG诱导组再分为两组,分别进行离心,弃去上清液,各加入1mL PBS重悬沉淀。再次离心弃去上清,各加入400μL PBS重悬沉淀。第一组取80μL作为IPTG诱导后的原液样品。第二组再重悬沉淀后再进行超声破碎,约三次,至菌液澄清即可。然后进行离心,收集80μL上清液作为IPTG诱导后的上清样。然后分离多余上清液,再加入100μL PBS重悬沉淀,取80μL样品,作为IPTG诱导后的沉淀样。
不进行IPTG诱导的对照组先进行离心,弃去上清后加入1mL PBS重悬沉淀,再次离心并弃去上清,加入400μL PBS重悬沉淀,然后取80μL作为对照样。
收集菌体,制备蛋白样,制备流程如图4。样品各加20μL 5×loading Buffer,吹打混均;进行100℃金属浴10 min;12000 rpm,离心10 min后,用于SDS-PAGE电泳检测。
重组菌表达鉴定结果如图5:SDS-PAGE检测诱导前后的两种重组菌体蛋白,结果显示,经过IPTG诱导的pET-16b-ErSpaA/BL21 star表达菌与未添加IPTG诱导的重组菌相比,在66kDa处有一条特异蛋白条带,与重组的SpaA蛋白的理论分子量66kDa相符。同理,经过IPTG诱导的pET-16b-ErSpaA/Rossetta gami B表达菌与未添加IPTG诱导的重组菌相比,在66kDa处有一条特异蛋白条带,同样与重组的SpaA蛋白的理论分子量66kDa相符。结果显示IPTG的诱导表达培养在室温下,摇床培养4小时便可达到最佳。
实施例4 重组蛋白ErSpaA的纯化
a. 大量培养重组菌,收集菌体
将获得的单克隆重组菌pET-16b-ErSpaA/Rossetta gami B接种100μL到5 mL含有Amp抗生素的LB液体培养基中,摇床培养过夜;第二天早上将上述菌液全部接种至1L LB液体培养基中,并加入10mg/mL的Amp 1 mL,在37℃,220 rpm条件下摇床培养6 h后,加入1 mLIPTG(1M)进行诱导;继续37℃,220 rpm摇床培养4h;将上述菌液用500 mL离心管分装成4管,每管250 mL,配平后,4000 rpm,离心30 min;弃去上清,每管加入30 mL PBS 重悬沉淀;准备4个50 mL 离心管,提前称重,将菌液转移到50 mL离心管中;将上述离心管配平,5000rpm,离心15 min,弃去上清;扣干,再次称重,得湿重;所得菌体于4 ℃冰箱保存。
进行所需试剂配置,配置比例如下:
A液 100 mL 1 M Tris-HCl(pH 8.0)+ 18 g NaCl,加1.9 L ddH2O 定容成2L;
B液(50 mM) 1.7 g咪唑+500 mL A液;
C液(200 mM)6.8 g咪唑+500 mL A液;
D液(500 mM)17 g 咪唑+500 mL A液。
b. 重组菌超声破碎
取细菌沉淀一管,加入20mL A液重悬,加1ml 10%SDS,然后进行超声破碎,程序设定为:On:3s,Off:5s,Ampl:80%,时长:15min,超声至澄清。将超声破碎后的细胞12000 rpm,4℃,离心10min,然后将上清转入新管放置冰上备用。
c. 重组蛋白亲和纯化
取3mL Ni2+填料悬浮液至层析柱,流干液体。加20mL去离子水,盖上盖子,颠倒5次,打开盖子流干液体,重复2次。加20 ml A液平衡Ni2+柱,颠倒混匀,打开盖子流干液体,重复2次。向Ni2+柱中加5 mL超声上清样品,颠倒混匀,冰上静置15min,期间每5分钟混合一次。打开盖子收集流穿液至新的离心管,标为“流穿”。加20ml A液洗涤,重复3次。加20ml B液洗涤,收中段样品1mL,标为“洗涤”。用2mL C液进行洗脱,收集样品,并标为“洗脱”,重复3次,分别收集。用20ml A液平衡Ni2+柱,重复3次。
继续上样5mL结合,重复以上步骤。
d. 蛋白超滤浓缩
样品进行SDS-PAGE凝胶电泳检测。将纯化得到的蛋白质样品用30 kD超滤管进行离心超滤浓缩,并用A液进行溶液置换,以降低溶液中的咪唑浓度。
蛋白质纯化结果如图6。
e. 重组蛋白的免疫实验:重组蛋白按照1:1与佐剂乳化(45 ug/200 ul),分2次间隔2周皮下注射小鼠,阴性对照注射生理盐水,免疫2周后对小鼠进行静脉采血,检测血清抗体。重组蛋白SpaA免疫小鼠后IgG含量与对照组相比均有所升高,OD450nm的测量数值和对照组分别为0.97和0.23,与对照组相比显著差异(p < 0.05)。由此可见,重组蛋白具有较好的免疫原性。
序列表
<110> 浙江理工大学
<120> 一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法
<130> 0000
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1734
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> misc_feature
<222> (1)..(1734)
<223> full gene sequence of mature ErSpaA
<400> 1
gacagcaccg atatcagcgt tattccgctg atcggtgaac aagttggtct gctgccggtg 60
ctgccgggta ccggcgtgca cgcgcaagag tacaacaaga tgaccgacgc gtatatcgaa 120
aaactggtta gcctgattaa ccagaaggtg aaaccgttcc tgatcaacga gccgaaaggt 180
taccaaagct ttgaagcggt taacgaggaa attaacagca tcgtgagcga gctgaagaac 240
gaaggcatga gcctgcagaa cattcaccac atgttcaaac agagcatcca aaacctggcg 300
acccgtattg gttaccgtag cttcatgcaa gatgcgatgt atctggagaa ctttgaacgt 360
ctgaccatcc cggagctgga cgaagcgtac gtggatctgc tggttaacta tgaggtgaag 420
caccgtatcc tggttaaata cgaaggcaag gtgaaaggcc gtgcgccgct ggaggcgttc 480
attgtgccgc tgcgtgaccg tatccgtagc atgaacgaga ttgcggcgga agttaactac 540
ctgccggagg cgcacgaaga ctttctggtt agcgatagca gcgaatacaa cgataagctg 600
aacaacatca acttcgcgct gggtctgggc gtgagcgagt ttattgacta caaccgtctg 660
gaaaacatga tggagaagga actgcacccg ctgtacctgg agctgtatgc gatgcgtcgt 720
aaccgtcaga tccaagtggt tcgtgatgtt tatccgaacc tggagcgtgc gaacgcggtg 780
gttgaaagcc tgaaaaccat taaggacatc aaacagcgtg gtaagaaact gcaagagctg 840
ctggaaatct acattcagcg tagcggtgac gttcgtaagc cggatgtgct gcagcgtttc 900
atcggcaaat atcaaagcgt ggttgatgag gaaaagaaca aactgcagga ctacctggaa 960
agcgacatct ttgatagcta tagcgtggat ggcgagaaga ttcgtaacaa agaaattacc 1020
ctgatcaacc gtgacgcgta cctgagcatg atttatcgtg cgcagagcat tagcgagatc 1080
aaaaccattc gtgcggatct ggaaagcctg gttaagagct tccaaaacga ggaaagcgac 1140
agcaaagtgg agccggaaag cccggtgaag gttgagaaac cggttgacaa ggaaaaaccg 1200
aaggatcaga agaaaccggt ggaccaaagc aagccggaga gcaacagcaa agaaggttgg 1260
atcaagaaag ataacaagtg gttctacatc gagaaaagcg gtggcatggc gaccggctgg 1320
aagaaagttg cggataagtg gtactatctg gacaacaccg gtgcgatcgt taccggctgg 1380
aagaaagtgg cgaacaagtg gtactatctg gagaagagcg gtgcgatggc gaccggttgg 1440
aagaaagtta gcaataaatg gtactatctg gagaacagcg gcgcgatggc tactggctgg 1500
aagaaagtga gcaacaaatg gtactatctg gaaaacagcg gcgcgatggc cactggctgg 1560
aagaaagtta gcaacaagtg gtattacctg gagaactctg gcgcgatggc tacaggttgg 1620
aagaaagttg cgaataaatg gtattacctg gacaaaagcg gtatgatggt gaccggcagc 1680
aagagcatcg atggtaagaa atacgcgttt aagaacgacg gcagcctgaa ataa 1734
<210> 2
<211> 597
<212> PRT
<213> 人工序列(Artificial Sequence)
<220>
<221> PEPTIDE
<222> (1)..(.597)
<223> ErSpaA antigen protein
<400> 2
Met Gly His His His His His His His His His His Ser Ser Gly His
1 5 10 15
Ile Glu Gly Arg Asp Ser Thr Asp Ile Ser Val Ile Pro Leu Ile Gly
20 25 30
Glu Gln Val Gly Leu Leu Pro Val Leu Pro Gly Thr Gly Val His Ala
35 40 45
Gln Glu Tyr Asn Lys Met Thr Asp Ala Tyr Ile Glu Lys Leu Val Ser
50 55 60
Leu Ile Asn Gln Lys Val Lys Pro Phe Leu Ile Asn Glu Pro Lys Gly
65 70 75 80
Tyr Gln Ser Phe Glu Ala Val Asn Glu Glu Ile Asn Ser Ile Val Ser
85 90 95
Glu Leu Lys Asn Glu Gly Met Ser Leu Gln Asn Ile His His Met Phe
100 105 110
Lys Gln Ser Ile Gln Asn Leu Ala Thr Arg Ile Gly Tyr Arg Ser Phe
115 120 125
Met Gln Asp Ala Met Tyr Leu Glu Asn Phe Glu Arg Leu Thr Ile Pro
130 135 140
Glu Leu Asp Glu Ala Tyr Val Asp Leu Leu Val Asn Tyr Glu Val Lys
145 150 155 160
His Arg Ile Leu Val Lys Tyr Glu Gly Lys Val Lys Gly Arg Ala Pro
165 170 175
Leu Glu Ala Phe Ile Val Pro Leu Arg Asp Arg Ile Arg Ser Met Asn
180 185 190
Glu Ile Ala Ala Glu Val Asn Tyr Leu Pro Glu Ala His Glu Asp Phe
195 200 205
Leu Val Ser Asp Ser Ser Glu Tyr Asn Asp Lys Leu Asn Asn Ile Asn
210 215 220
Phe Ala Leu Gly Leu Gly Val Ser Glu Phe Ile Asp Tyr Asn Arg Leu
225 230 235 240
Glu Asn Met Met Glu Lys Glu Leu His Pro Leu Tyr Leu Glu Leu Tyr
245 250 255
Ala Met Arg Arg Asn Arg Gln Ile Gln Val Val Arg Asp Val Tyr Pro
260 265 270
Asn Leu Glu Arg Ala Asn Ala Val Val Glu Ser Leu Lys Thr Ile Lys
275 280 285
Asp Ile Lys Gln Arg Gly Lys Lys Leu Gln Glu Leu Leu Glu Ile Tyr
290 295 300
Ile Gln Arg Ser Gly Asp Val Arg Lys Pro Asp Val Leu Gln Arg Phe
305 310 315 320
Ile Gly Lys Tyr Gln Ser Val Val Asp Glu Glu Lys Asn Lys Leu Gln
325 330 335
Asp Tyr Leu Glu Ser Asp Ile Phe Asp Ser Tyr Ser Val Asp Gly Glu
340 345 350
Lys Ile Arg Asn Lys Glu Ile Thr Leu Ile Asn Arg Asp Ala Tyr Leu
355 360 365
Ser Met Ile Tyr Arg Ala Gln Ser Ile Ser Glu Ile Lys Thr Ile Arg
370 375 380
Ala Asp Leu Glu Ser Leu Val Lys Ser Phe Gln Asn Glu Glu Ser Asp
385 390 395 400
Ser Lys Val Glu Pro Glu Ser Pro Val Lys Val Glu Lys Pro Val Asp
405 410 415
Lys Glu Lys Pro Lys Asp Gln Lys Lys Pro Val Asp Gln Ser Lys Pro
420 425 430
Glu Ser Asn Ser Lys Glu Gly Trp Ile Lys Lys Asp Asn Lys Trp Phe
435 440 445
Tyr Ile Glu Lys Ser Gly Gly Met Ala Thr Gly Trp Lys Lys Val Ala
450 455 460
Asp Lys Trp Tyr Tyr Leu Asp Asn Thr Gly Ala Ile Val Thr Gly Trp
465 470 475 480
Lys Lys Val Ala Asn Lys Trp Tyr Tyr Leu Glu Lys Ser Gly Ala Met
485 490 495
Ala Thr Gly Trp Lys Lys Val Ser Asn Lys Trp Tyr Tyr Leu Glu Asn
500 505 510
Ser Gly Ala Met Ala Thr Gly Trp Lys Lys Val Ser Asn Lys Trp Tyr
515 520 525
Tyr Leu Glu Asn Ser Gly Ala Met Ala Thr Gly Trp Lys Lys Val Ser
530 535 540
Asn Lys Trp Tyr Tyr Leu Glu Asn Ser Gly Ala Met Ala Thr Gly Trp
545 550 555 560
Lys Lys Val Ala Asn Lys Trp Tyr Tyr Leu Asp Lys Ser Gly Met Met
565 570 575
Val Thr Gly Ser Lys Ser Ile Asp Gly Lys Lys Tyr Ala Phe Lys Asn
580 585 590
Asp Gly Ser Leu Lys
595
Claims (10)
1.一种猪丹毒SpaA抗原蛋白,其特征在于,其编码基因序列与SEQ ID NO.1具有大于90%的同一性,或大于92%的同一性,或大于95%的同一性,或大于96%的同一性,或大于97%的同一性,或大于98%的同一性,或大于99%的同一性;优选如SEQ ID NO.1所示。
2.一种猪丹毒SpaA抗原蛋白,其特征在于,其氨基酸序列与SEQ ID NO.2具有大于90%的同一性,或大于92%的同一性,或大于95%的同一性,或大于96%的同一性,或大于97%的同一性,或大于98%的同一性,或大于99%的同一性;优选如SEQ ID NO.2所示。
3.一种重组载体,其特征在于,含有权利要求1或2所述的猪丹毒SpaA抗原蛋白的编码基因;优选是pET-16b-ErSpaA。
4.一种猪丹毒SpaA基因工程疫苗,其特征在于,所述疫苗包含权利要求1或2所述的猪丹毒SpaA抗原蛋白;或包含含有权利要求3所述的重组载体。
5.根据权利要求4所述的猪丹毒SpaA基因工程疫苗,其特征在于,还包含疫苗佐剂、助悬剂、表面活性剂、抗原灭活剂和/或防腐剂。
6.如权利要求1或2所述的猪丹毒SpaA抗原蛋白优化克隆的制备方法,其特征在于,包括如下步骤:
构建含有上述重组蛋白基因的表达载体;
重组蛋白的表达。
7.根据权利要求6所述的制备方法,其特征在于,还包括对表达系统表达重组蛋白的纯化的步骤;
任选地,还包括在构建表达载体前对目的基因的获取步骤。
8.根据权利要求7所述的制备方法,其特征在于,所述制备方法的具体步骤为:
将获取的目的基因酶切后连接入pET-16b载体,转入克隆菌,获得重组载体pET-16b-ErSpaA;
上述重组载体分别转入表达菌中,诱导表达目的蛋白。
9.根据权利要求8所述的制备方法,其特征在于,所述克隆菌优选甘油菌,表达菌优选Rossetta gami B或BL21 star。
10.如权利要求1或2所述的猪丹毒SpaA抗原蛋白、权利要求3所述的重组载体、权利要求4所述的基因工程疫苗在制备抗猪丹毒杆菌感染的疫苗中的应用。
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202010991427 | 2020-09-21 | ||
CN2020109914278 | 2020-09-21 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN113388624A true CN113388624A (zh) | 2021-09-14 |
Family
ID=77616600
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202011366261.7A Pending CN113388624A (zh) | 2020-09-21 | 2020-11-29 | 一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN113388624A (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN115850404A (zh) * | 2022-12-13 | 2023-03-28 | 中国兽医药品监察所 | 一种串联优势表位的重组猪丹毒杆菌表面保护抗原a及其应用 |
Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2000047744A1 (en) * | 1999-02-10 | 2000-08-17 | The Rockefeller University | Antigen of erysipelothirx rhusiopathiae comprising an immuno-protective epitope |
CN101031647A (zh) * | 2004-02-27 | 2007-09-05 | 财团法人化学及血清疗法研究所 | 在大肠杆菌中制备猪丹毒杆菌表面保护性抗原变体的方法 |
CN106146626A (zh) * | 2015-04-07 | 2016-11-23 | 武汉科缘生物发展有限责任公司 | 一种红斑丹毒丝菌亚单位疫苗及制备方法及应用 |
TWI638827B (zh) * | 2017-07-25 | 2018-10-21 | 國立屏東科技大學 | 交叉保護性重組抗原及含彼之動物疫苗組成物 |
CN110183520A (zh) * | 2019-05-25 | 2019-08-30 | 青岛易邦生物工程有限公司 | 一种猪丹毒SpaA蛋白及其在制备疫苗中的应用 |
-
2020
- 2020-11-29 CN CN202011366261.7A patent/CN113388624A/zh active Pending
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2000047744A1 (en) * | 1999-02-10 | 2000-08-17 | The Rockefeller University | Antigen of erysipelothirx rhusiopathiae comprising an immuno-protective epitope |
CN101031647A (zh) * | 2004-02-27 | 2007-09-05 | 财团法人化学及血清疗法研究所 | 在大肠杆菌中制备猪丹毒杆菌表面保护性抗原变体的方法 |
CN106146626A (zh) * | 2015-04-07 | 2016-11-23 | 武汉科缘生物发展有限责任公司 | 一种红斑丹毒丝菌亚单位疫苗及制备方法及应用 |
TWI638827B (zh) * | 2017-07-25 | 2018-10-21 | 國立屏東科技大學 | 交叉保護性重組抗原及含彼之動物疫苗組成物 |
CN110183520A (zh) * | 2019-05-25 | 2019-08-30 | 青岛易邦生物工程有限公司 | 一种猪丹毒SpaA蛋白及其在制备疫苗中的应用 |
Non-Patent Citations (1)
Title |
---|
MAKINO S等: "《Properties of repeat domain found in a novel protective antigen, SpaA, of Erysipelothrix rhusiopathiae》", 《MICROBIAL PATHOGENESIS》 * |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN115850404A (zh) * | 2022-12-13 | 2023-03-28 | 中国兽医药品监察所 | 一种串联优势表位的重组猪丹毒杆菌表面保护抗原a及其应用 |
CN115850404B (zh) * | 2022-12-13 | 2024-02-09 | 中国兽医药品监察所 | 一种串联优势表位的重组猪丹毒杆菌表面保护抗原a及其应用 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN113150171B (zh) | 含分子内佐剂的非洲猪瘟病毒重组蛋白、表达载体及应用 | |
US7951386B2 (en) | Ehrlichia canis genes and vaccines | |
CN110183520B (zh) | 一种猪丹毒SpaA蛋白及其在制备疫苗中的应用 | |
CN109867727B (zh) | 一种flagellin-fiber2融合蛋白、其制备方法和应用 | |
CN111856006B (zh) | 牛支原体分泌蛋白MbovP274的应用 | |
JPH11501809A (ja) | マイコプラズマに対する抗原組成物 | |
CN113943376B (zh) | 一种融合基因、其编码蛋白及其在抗非洲猪瘟中的应用 | |
CN109207502B (zh) | 猪支原体肺炎和猪圆环病毒2型重组蛋白及制备二联疫苗 | |
CN108330142B (zh) | 一种具有免疫保护作用的美人鱼发光杆菌溶血素Hlych蛋白 | |
CN113388624A (zh) | 一种猪丹毒SpaA抗原蛋白及其优化克隆的制备方法 | |
CN112159479B (zh) | 一种鸡毒支原体多抗原表位融合蛋白pMG-mEA及其应用 | |
CN105646681B (zh) | 一种奶牛金黄色葡萄球菌α-溶血素亚单位疫苗的制备方法及应用 | |
CN115850404B (zh) | 一种串联优势表位的重组猪丹毒杆菌表面保护抗原a及其应用 | |
CN110746496A (zh) | 一种鲍曼不动杆菌的pal重组蛋白及其编码基因和它们的应用 | |
CN112279925B (zh) | 一种融合蛋白及一种犬弓形虫亚单位疫苗及其疫苗组合物 | |
CN104508120A (zh) | 编码肝素结合血凝素(hbha)融合蛋白质的重组分枝杆菌和其用途 | |
CN114903986A (zh) | 一种猪链球菌三组分亚单位疫苗其制备方法 | |
CN111138553B (zh) | 一种融合蛋白及一种弓形虫亚单位疫苗及其疫苗组合物 | |
CN114437236A (zh) | 一种重组非洲猪瘟病毒多表位融合蛋白、制备及其应用 | |
CN107502616B (zh) | 一种可溶性重组蛋白cta-cd154及其制备方法和应用 | |
CN114058634A (zh) | 一种鸡滑液囊支原体基因工程亚单位疫苗 | |
CN114456240B (zh) | 一种非洲猪瘟病毒基因工程亚单位口服疫苗 | |
Basiri et al. | Amplification, cloning and expression of Brucella melitensis bp26 gene (OMP28) isolated from Markazi province (Iran) and purification of Bp26 Protein | |
RU2590701C2 (ru) | Антигенная композиция и ее терапевтическое применение для профилактики и лечения онкологических заболеваний, рекомбинантная плазмидная днк, обеспечивающая синтез гибридного белка, а также способ получения белка | |
CN114073762A (zh) | 一种含有Omp19和VirB8蛋白的布鲁氏菌病双组分疫苗 |
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 | ||
RJ01 | Rejection of invention patent application after publication | ||
RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210914 |