CN113214389A - 全人源的新冠IgL单链抗体及其应用 - Google Patents
全人源的新冠IgL单链抗体及其应用 Download PDFInfo
- Publication number
- CN113214389A CN113214389A CN202110545795.4A CN202110545795A CN113214389A CN 113214389 A CN113214389 A CN 113214389A CN 202110545795 A CN202110545795 A CN 202110545795A CN 113214389 A CN113214389 A CN 113214389A
- Authority
- CN
- China
- Prior art keywords
- antibody
- bcr
- sequence
- igl
- ser
- 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
- 238000012163 sequencing technique Methods 0.000 claims abstract description 54
- 101000933320 Homo sapiens Breakpoint cluster region protein Proteins 0.000 claims abstract description 10
- 238000001514 detection method Methods 0.000 claims abstract description 8
- 238000002360 preparation method Methods 0.000 claims abstract description 6
- 108090000623 proteins and genes Proteins 0.000 claims description 28
- 238000000034 method Methods 0.000 claims description 18
- 150000001413 amino acids Chemical class 0.000 claims description 13
- 241000711573 Coronaviridae Species 0.000 claims description 12
- 238000012216 screening Methods 0.000 claims description 11
- 239000013604 expression vector Substances 0.000 claims description 8
- 239000002773 nucleotide Substances 0.000 claims description 8
- 125000003729 nucleotide group Chemical group 0.000 claims description 8
- 230000008569 process Effects 0.000 claims description 7
- 238000012937 correction Methods 0.000 claims description 4
- 238000004458 analytical method Methods 0.000 claims description 3
- 239000003937 drug carrier Substances 0.000 claims description 2
- 150000003839 salts Chemical class 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 229940126585 therapeutic drug Drugs 0.000 claims 1
- 230000003321 amplification Effects 0.000 abstract description 35
- 238000003199 nucleic acid amplification method Methods 0.000 abstract description 35
- 238000004519 manufacturing process Methods 0.000 abstract description 8
- 229960005486 vaccine Drugs 0.000 abstract description 7
- 241000700605 Viruses Species 0.000 abstract description 5
- 238000011161 development Methods 0.000 abstract description 5
- 238000009509 drug development Methods 0.000 abstract description 5
- 229940079593 drug Drugs 0.000 abstract description 4
- 239000003814 drug Substances 0.000 abstract description 4
- 238000010353 genetic engineering Methods 0.000 abstract description 4
- 239000003550 marker Substances 0.000 abstract description 4
- 238000013373 clone screening Methods 0.000 abstract 1
- 230000003472 neutralizing effect Effects 0.000 abstract 1
- 108091008875 B cell receptors Proteins 0.000 description 76
- 238000006243 chemical reaction Methods 0.000 description 21
- 210000004027 cell Anatomy 0.000 description 13
- 239000012634 fragment Substances 0.000 description 13
- 108020004414 DNA Proteins 0.000 description 12
- 239000000523 sample Substances 0.000 description 12
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 12
- 238000002156 mixing Methods 0.000 description 11
- 108060003951 Immunoglobulin Proteins 0.000 description 10
- 210000003719 b-lymphocyte Anatomy 0.000 description 10
- 102000018358 immunoglobulin Human genes 0.000 description 10
- 239000000047 product Substances 0.000 description 10
- 239000002299 complementary DNA Substances 0.000 description 8
- 238000013519 translation Methods 0.000 description 8
- 108020004705 Codon Proteins 0.000 description 7
- 230000014509 gene expression Effects 0.000 description 7
- 102000004169 proteins and genes Human genes 0.000 description 7
- 108091028043 Nucleic acid sequence Proteins 0.000 description 6
- 239000011324 bead Substances 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 108091093088 Amplicon Proteins 0.000 description 5
- 239000003153 chemical reaction reagent Substances 0.000 description 5
- 238000001962 electrophoresis Methods 0.000 description 5
- 229940072221 immunoglobulins Drugs 0.000 description 5
- 238000012408 PCR amplification Methods 0.000 description 4
- UIGMAMGZOJVTDN-WHFBIAKZSA-N Ser-Gly-Ser Chemical compound OC[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O UIGMAMGZOJVTDN-WHFBIAKZSA-N 0.000 description 4
- 108091081024 Start codon Proteins 0.000 description 4
- 239000000427 antigen Substances 0.000 description 4
- 102000036639 antigens Human genes 0.000 description 4
- 108091007433 antigens Proteins 0.000 description 4
- 238000005119 centrifugation Methods 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 108010025153 lysyl-alanyl-alanine Proteins 0.000 description 4
- 108010031719 prolyl-serine Proteins 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 3
- VAWNQIGQPUOPQW-ACZMJKKPSA-N Asp-Glu-Ala Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(O)=O VAWNQIGQPUOPQW-ACZMJKKPSA-N 0.000 description 3
- ALMIMUZAWTUNIO-BZSNNMDCSA-N Asp-Tyr-Tyr 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](CC1=CC=C(O)C=C1)C(O)=O ALMIMUZAWTUNIO-BZSNNMDCSA-N 0.000 description 3
- 102000019260 B-Cell Antigen Receptors Human genes 0.000 description 3
- 108010012919 B-Cell Antigen Receptors Proteins 0.000 description 3
- FBGXMKUWQFPHFB-JBDRJPRFSA-N Ile-Ser-Cys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CS)C(=O)O)N FBGXMKUWQFPHFB-JBDRJPRFSA-N 0.000 description 3
- JMVQDLDPDBXAAX-YUMQZZPRSA-N Pro-Gly-Gln Chemical compound NC(=O)CC[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H]1CCCN1 JMVQDLDPDBXAAX-YUMQZZPRSA-N 0.000 description 3
- SIEBDTCABMZCLF-XGEHTFHBSA-N Ser-Val-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O SIEBDTCABMZCLF-XGEHTFHBSA-N 0.000 description 3
- 108020005038 Terminator Codon Proteins 0.000 description 3
- ZLFHAAGHGQBQQN-GUBZILKMSA-N Val-Ala-Pro Natural products CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(O)=O ZLFHAAGHGQBQQN-GUBZILKMSA-N 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 238000010276 construction Methods 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 239000012149 elution buffer Substances 0.000 description 3
- 108010027668 glycyl-alanyl-valine Proteins 0.000 description 3
- XKUKSGPZAADMRA-UHFFFAOYSA-N glycyl-glycyl-glycine Natural products NCC(=O)NCC(=O)NCC(O)=O XKUKSGPZAADMRA-UHFFFAOYSA-N 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 108010070409 phenylalanyl-glycyl-glycine Proteins 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 239000002096 quantum dot Substances 0.000 description 3
- 230000008707 rearrangement Effects 0.000 description 3
- 238000013518 transcription Methods 0.000 description 3
- 230000035897 transcription Effects 0.000 description 3
- CXRCVCURMBFFOL-FXQIFTODSA-N Ala-Ala-Pro Chemical compound C[C@H](N)C(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(O)=O CXRCVCURMBFFOL-FXQIFTODSA-N 0.000 description 2
- 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
- FFZJHQODAYHGPO-KZVJFYERSA-N Ala-Pro-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@H](C)N FFZJHQODAYHGPO-KZVJFYERSA-N 0.000 description 2
- IOFVWPYSRSCWHI-JXUBOQSCSA-N Ala-Thr-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](C)N IOFVWPYSRSCWHI-JXUBOQSCSA-N 0.000 description 2
- OMSKGWFGWCQFBD-KZVJFYERSA-N Ala-Val-Thr Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O OMSKGWFGWCQFBD-KZVJFYERSA-N 0.000 description 2
- 101710095183 B-cell antigen receptor complex-associated protein alpha chain Proteins 0.000 description 2
- 102100027205 B-cell antigen receptor complex-associated protein alpha chain Human genes 0.000 description 2
- 101710166261 B-cell antigen receptor complex-associated protein beta chain Proteins 0.000 description 2
- 102100027203 B-cell antigen receptor complex-associated protein beta chain Human genes 0.000 description 2
- 101150049556 Bcr gene Proteins 0.000 description 2
- 241001678559 COVID-19 virus Species 0.000 description 2
- SFRQEQGPRTVDPO-NRPADANISA-N Cys-Gln-Val Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(O)=O SFRQEQGPRTVDPO-NRPADANISA-N 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 241000588724 Escherichia coli Species 0.000 description 2
- RZSLYUUFFVHFRQ-FXQIFTODSA-N Gln-Ala-Glu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O RZSLYUUFFVHFRQ-FXQIFTODSA-N 0.000 description 2
- MFORDNZDKAVNSR-SRVKXCTJSA-N Gln-Pro-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)CCC(N)=O MFORDNZDKAVNSR-SRVKXCTJSA-N 0.000 description 2
- BETSEXMYBWCDAE-SZMVWBNQSA-N Gln-Trp-Lys Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)N)N BETSEXMYBWCDAE-SZMVWBNQSA-N 0.000 description 2
- HNAUFGBKJLTWQE-IFFSRLJSSA-N Gln-Val-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CCC(=O)N)N)O HNAUFGBKJLTWQE-IFFSRLJSSA-N 0.000 description 2
- KVBPDJIFRQUQFY-ACZMJKKPSA-N Glu-Cys-Ser Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CS)C(=O)N[C@@H](CO)C(O)=O KVBPDJIFRQUQFY-ACZMJKKPSA-N 0.000 description 2
- DNPCBMNFQVTHMA-DCAQKATOSA-N Glu-Leu-Gln Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O DNPCBMNFQVTHMA-DCAQKATOSA-N 0.000 description 2
- UQJNXZSSGQIPIQ-FBCQKBJTSA-N Gly-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)CN UQJNXZSSGQIPIQ-FBCQKBJTSA-N 0.000 description 2
- 241000238631 Hexapoda Species 0.000 description 2
- ZPVJJPAIUZLSNE-DCAQKATOSA-N His-Arg-Ser Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(O)=O ZPVJJPAIUZLSNE-DCAQKATOSA-N 0.000 description 2
- AKEDPWJFQULLPE-IUCAKERBSA-N His-Glu-Gly Chemical compound N[C@@H](Cc1cnc[nH]1)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O AKEDPWJFQULLPE-IUCAKERBSA-N 0.000 description 2
- FOCSWPCHUDVNLP-PMVMPFDFSA-N His-Trp-Tyr Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CC3=CC=C(C=C3)O)C(=O)O)NC(=O)[C@H](CC4=CN=CN4)N FOCSWPCHUDVNLP-PMVMPFDFSA-N 0.000 description 2
- JZNVOBUNTWNZPW-GHCJXIJMSA-N Ile-Ser-Asp Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)O)C(=O)O)N JZNVOBUNTWNZPW-GHCJXIJMSA-N 0.000 description 2
- 102000013463 Immunoglobulin Light Chains Human genes 0.000 description 2
- 108010065825 Immunoglobulin Light Chains Proteins 0.000 description 2
- 239000012880 LB liquid culture medium Substances 0.000 description 2
- 241000880493 Leptailurus serval Species 0.000 description 2
- UHNQRAFSEBGZFZ-YESZJQIVSA-N Leu-Phe-Pro Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N2CCC[C@@H]2C(=O)O)N UHNQRAFSEBGZFZ-YESZJQIVSA-N 0.000 description 2
- XOWMDXHFSBCAKQ-SRVKXCTJSA-N Leu-Ser-Leu Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC(C)C XOWMDXHFSBCAKQ-SRVKXCTJSA-N 0.000 description 2
- LCNASHSOFMRYFO-WDCWCFNPSA-N Leu-Thr-Gln Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@H](C(O)=O)CCC(N)=O LCNASHSOFMRYFO-WDCWCFNPSA-N 0.000 description 2
- FZIJIFCXUCZHOL-CIUDSAMLSA-N Lys-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN FZIJIFCXUCZHOL-CIUDSAMLSA-N 0.000 description 2
- MPGHETGWWWUHPY-CIUDSAMLSA-N Lys-Ala-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN MPGHETGWWWUHPY-CIUDSAMLSA-N 0.000 description 2
- XFIHDSBIPWEYJJ-YUMQZZPRSA-N Lys-Ala-Gly Chemical compound OC(=O)CNC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN XFIHDSBIPWEYJJ-YUMQZZPRSA-N 0.000 description 2
- FUKDBQGFSJUXGX-RWMBFGLXSA-N Lys-Arg-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CCCCN)N)C(=O)O FUKDBQGFSJUXGX-RWMBFGLXSA-N 0.000 description 2
- OIQSIMFSVLLWBX-VOAKCMCISA-N Lys-Leu-Thr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O OIQSIMFSVLLWBX-VOAKCMCISA-N 0.000 description 2
- 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 2
- VHTOGMKQXXJOHG-RHYQMDGZSA-N Lys-Thr-Val Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O VHTOGMKQXXJOHG-RHYQMDGZSA-N 0.000 description 2
- 241000127282 Middle East respiratory syndrome-related coronavirus Species 0.000 description 2
- 102100030569 Nuclear receptor corepressor 2 Human genes 0.000 description 2
- 101710153660 Nuclear receptor corepressor 2 Proteins 0.000 description 2
- UIMCLYYSUCIUJM-UWVGGRQHSA-N Pro-Gly-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H]1CCCN1 UIMCLYYSUCIUJM-UWVGGRQHSA-N 0.000 description 2
- RMODQFBNDDENCP-IHRRRGAJSA-N Pro-Lys-Leu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O RMODQFBNDDENCP-IHRRRGAJSA-N 0.000 description 2
- FDMKYQQYJKYCLV-GUBZILKMSA-N Pro-Pro-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@H]1NCCC1 FDMKYQQYJKYCLV-GUBZILKMSA-N 0.000 description 2
- MKGIILKDUGDRRO-FXQIFTODSA-N Pro-Ser-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H]1CCCN1 MKGIILKDUGDRRO-FXQIFTODSA-N 0.000 description 2
- 206010037660 Pyrexia Diseases 0.000 description 2
- 241000315672 SARS coronavirus Species 0.000 description 2
- OOKCGAYXSNJBGQ-ZLUOBGJFSA-N Ser-Asn-Asn Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O OOKCGAYXSNJBGQ-ZLUOBGJFSA-N 0.000 description 2
- IOVHBRCQOGWAQH-ZKWXMUAHSA-N Ser-Gly-Ile Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H]([C@@H](C)CC)C(O)=O IOVHBRCQOGWAQH-ZKWXMUAHSA-N 0.000 description 2
- BYCVMHKULKRVPV-GUBZILKMSA-N Ser-Lys-Gln Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(N)=O)C(O)=O BYCVMHKULKRVPV-GUBZILKMSA-N 0.000 description 2
- CUXJENOFJXOSOZ-BIIVOSGPSA-N Ser-Ser-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CO)NC(=O)[C@H](CO)N)C(=O)O CUXJENOFJXOSOZ-BIIVOSGPSA-N 0.000 description 2
- OLKICIBQRVSQMA-SRVKXCTJSA-N Ser-Ser-Tyr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O OLKICIBQRVSQMA-SRVKXCTJSA-N 0.000 description 2
- BDMWLJLPPUCLNV-XGEHTFHBSA-N Ser-Thr-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O BDMWLJLPPUCLNV-XGEHTFHBSA-N 0.000 description 2
- STIAINRLUUKYKM-WFBYXXMGSA-N Ser-Trp-Ala Chemical compound C1=CC=C2C(C[C@@H](C(=O)N[C@@H](C)C(O)=O)NC(=O)[C@@H](N)CO)=CNC2=C1 STIAINRLUUKYKM-WFBYXXMGSA-N 0.000 description 2
- KBBRNEDOYWMIJP-KYNKHSRBSA-N Thr-Gly-Thr Chemical compound C[C@H]([C@@H](C(=O)NCC(=O)N[C@@H]([C@@H](C)O)C(=O)O)N)O KBBRNEDOYWMIJP-KYNKHSRBSA-N 0.000 description 2
- XKWABWFMQXMUMT-HJGDQZAQSA-N Thr-Pro-Glu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(O)=O XKWABWFMQXMUMT-HJGDQZAQSA-N 0.000 description 2
- ZESGVALRVJIVLZ-VFCFLDTKSA-N Thr-Thr-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H]([C@@H](C)O)C(=O)N1CCC[C@@H]1C(=O)O)N)O ZESGVALRVJIVLZ-VFCFLDTKSA-N 0.000 description 2
- VCXWRWYFJLXITF-AUTRQRHGSA-N Tyr-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC1=CC=C(O)C=C1 VCXWRWYFJLXITF-AUTRQRHGSA-N 0.000 description 2
- SZEIFUXUTBBQFQ-STQMWFEESA-N Tyr-Pro-Gly Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O SZEIFUXUTBBQFQ-STQMWFEESA-N 0.000 description 2
- KWKJGBHDYJOVCR-SRVKXCTJSA-N Tyr-Ser-Cys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CS)C(=O)O)N)O KWKJGBHDYJOVCR-SRVKXCTJSA-N 0.000 description 2
- VJOWWOGRNXRQMF-UVBJJODRSA-N Val-Ala-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@H](C)NC(=O)[C@@H](N)C(C)C)C(O)=O)=CNC2=C1 VJOWWOGRNXRQMF-UVBJJODRSA-N 0.000 description 2
- LHADRQBREKTRLR-DCAQKATOSA-N Val-Cys-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](C(C)C)N LHADRQBREKTRLR-DCAQKATOSA-N 0.000 description 2
- XWYUBUYQMOUFRQ-IFFSRLJSSA-N Val-Glu-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](C(C)C)N)O XWYUBUYQMOUFRQ-IFFSRLJSSA-N 0.000 description 2
- UMPVMAYCLYMYGA-ONGXEEELSA-N Val-Leu-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O UMPVMAYCLYMYGA-ONGXEEELSA-N 0.000 description 2
- CKTMJBPRVQWPHU-JSGCOSHPSA-N Val-Phe-Gly Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)NCC(=O)O)N CKTMJBPRVQWPHU-JSGCOSHPSA-N 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- 108010008685 alanyl-glutamyl-aspartic acid Proteins 0.000 description 2
- 108010044940 alanylglutamine Proteins 0.000 description 2
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 2
- 229960000723 ampicillin Drugs 0.000 description 2
- 229940125644 antibody drug Drugs 0.000 description 2
- 108010069205 aspartyl-phenylalanine Proteins 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 210000000349 chromosome Anatomy 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 102000051878 human BCR Human genes 0.000 description 2
- 230000008105 immune reaction Effects 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 230000003834 intracellular effect Effects 0.000 description 2
- 210000000265 leukocyte Anatomy 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 239000013610 patient sample Substances 0.000 description 2
- 108010051242 phenylalanylserine Proteins 0.000 description 2
- 229920001184 polypeptide Polymers 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 102000004196 processed proteins & peptides Human genes 0.000 description 2
- 230000009465 prokaryotic expression Effects 0.000 description 2
- 108010077112 prolyl-proline Proteins 0.000 description 2
- 238000011002 quantification Methods 0.000 description 2
- 230000008439 repair process Effects 0.000 description 2
- 238000012827 research and development Methods 0.000 description 2
- 238000010839 reverse transcription Methods 0.000 description 2
- 238000005096 rolling process Methods 0.000 description 2
- 230000019491 signal transduction Effects 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 239000013598 vector Substances 0.000 description 2
- 238000012795 verification Methods 0.000 description 2
- 230000009385 viral infection Effects 0.000 description 2
- 208000010444 Acidosis Diseases 0.000 description 1
- 206010001052 Acute respiratory distress syndrome Diseases 0.000 description 1
- NBTGEURICRTMGL-WHFBIAKZSA-N Ala-Gly-Ser Chemical compound C[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O NBTGEURICRTMGL-WHFBIAKZSA-N 0.000 description 1
- YHBDGLZYNIARKJ-GUBZILKMSA-N Ala-Pro-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@H](C)N YHBDGLZYNIARKJ-GUBZILKMSA-N 0.000 description 1
- NCQMBSJGJMYKCK-ZLUOBGJFSA-N Ala-Ser-Ser Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O NCQMBSJGJMYKCK-ZLUOBGJFSA-N 0.000 description 1
- ARHJJAAWNWOACN-FXQIFTODSA-N Ala-Ser-Val Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O ARHJJAAWNWOACN-FXQIFTODSA-N 0.000 description 1
- 102000006306 Antigen Receptors Human genes 0.000 description 1
- 108010083359 Antigen Receptors Proteins 0.000 description 1
- QLSRIZIDQXDQHK-RCWTZXSCSA-N Arg-Val-Thr Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O QLSRIZIDQXDQHK-RCWTZXSCSA-N 0.000 description 1
- HUZGPXBILPMCHM-IHRRRGAJSA-N Asn-Arg-Phe Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O HUZGPXBILPMCHM-IHRRRGAJSA-N 0.000 description 1
- APHUDFFMXFYRKP-CIUDSAMLSA-N Asn-Asn-Lys Chemical compound C(CCN)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](CC(=O)N)N APHUDFFMXFYRKP-CIUDSAMLSA-N 0.000 description 1
- NVWJMQNYLYWVNQ-BYULHYEWSA-N Asn-Ile-Gly Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(O)=O NVWJMQNYLYWVNQ-BYULHYEWSA-N 0.000 description 1
- NCFJQJRLQJEECD-NHCYSSNCSA-N Asn-Leu-Val Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(O)=O NCFJQJRLQJEECD-NHCYSSNCSA-N 0.000 description 1
- XTMZYFMTYJNABC-ZLUOBGJFSA-N Asn-Ser-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(=O)N)N XTMZYFMTYJNABC-ZLUOBGJFSA-N 0.000 description 1
- WLVLIYYBPPONRJ-GCJQMDKQSA-N Asn-Thr-Ala Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O WLVLIYYBPPONRJ-GCJQMDKQSA-N 0.000 description 1
- DBWYWXNMZZYIRY-LPEHRKFASA-N Asp-Arg-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(=O)O)N)C(=O)O DBWYWXNMZZYIRY-LPEHRKFASA-N 0.000 description 1
- XXAMCEGRCZQGEM-ZLUOBGJFSA-N Asp-Ser-Asn Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O XXAMCEGRCZQGEM-ZLUOBGJFSA-N 0.000 description 1
- 230000003844 B-cell-activation Effects 0.000 description 1
- 241000288673 Chiroptera Species 0.000 description 1
- 102100031673 Corneodesmosin Human genes 0.000 description 1
- 101710139375 Corneodesmosin Proteins 0.000 description 1
- 206010011224 Cough Diseases 0.000 description 1
- ATPDEYTYWVMINF-ZLUOBGJFSA-N Cys-Cys-Ser Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CS)C(=O)N[C@@H](CO)C(O)=O ATPDEYTYWVMINF-ZLUOBGJFSA-N 0.000 description 1
- HHABWQIFXZPZCK-ACZMJKKPSA-N Cys-Gln-Ser Chemical compound C(CC(=O)N)[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CS)N HHABWQIFXZPZCK-ACZMJKKPSA-N 0.000 description 1
- 101100193633 Danio rerio rag2 gene Proteins 0.000 description 1
- 206010012735 Diarrhoea Diseases 0.000 description 1
- 101710091045 Envelope protein Proteins 0.000 description 1
- 101710204837 Envelope small membrane protein Proteins 0.000 description 1
- 241000195955 Equisetum hyemale Species 0.000 description 1
- 108060002716 Exonuclease Proteins 0.000 description 1
- WUAYFMZULZDSLB-ACZMJKKPSA-N Gln-Ala-Asn Chemical compound NC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCC(N)=O WUAYFMZULZDSLB-ACZMJKKPSA-N 0.000 description 1
- GPISLLFQNHELLK-DCAQKATOSA-N Gln-Gln-His Chemical compound C1=C(NC=N1)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCC(=O)N)N GPISLLFQNHELLK-DCAQKATOSA-N 0.000 description 1
- KVXVVDFOZNYYKZ-DCAQKATOSA-N Gln-Gln-Leu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(O)=O KVXVVDFOZNYYKZ-DCAQKATOSA-N 0.000 description 1
- MADFVRSKEIEZHZ-DCAQKATOSA-N Gln-Gln-Lys Chemical compound C(CCN)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCC(=O)N)N MADFVRSKEIEZHZ-DCAQKATOSA-N 0.000 description 1
- KUBFPYIMAGXGBT-ACZMJKKPSA-N Gln-Ser-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(O)=O KUBFPYIMAGXGBT-ACZMJKKPSA-N 0.000 description 1
- OTQSTOXRUBVWAP-NRPADANISA-N Gln-Ser-Val Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O OTQSTOXRUBVWAP-NRPADANISA-N 0.000 description 1
- UTKUTMJSWKKHEM-WDSKDSINSA-N Glu-Ala-Gly Chemical compound OC(=O)CNC(=O)[C@H](C)NC(=O)[C@@H](N)CCC(O)=O UTKUTMJSWKKHEM-WDSKDSINSA-N 0.000 description 1
- MUSGDMDGNGXULI-DCAQKATOSA-N Glu-Glu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CCC(O)=O MUSGDMDGNGXULI-DCAQKATOSA-N 0.000 description 1
- RAUDKMVXNOWDLS-WDSKDSINSA-N Glu-Gly-Ser Chemical compound OC(=O)CC[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O RAUDKMVXNOWDLS-WDSKDSINSA-N 0.000 description 1
- QDMVXRNLOPTPIE-WDCWCFNPSA-N Glu-Lys-Thr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(O)=O QDMVXRNLOPTPIE-WDCWCFNPSA-N 0.000 description 1
- UGVQELHRNUDMAA-BYPYZUCNSA-N Gly-Ala-Gly Chemical compound [NH3+]CC(=O)N[C@@H](C)C(=O)NCC([O-])=O UGVQELHRNUDMAA-BYPYZUCNSA-N 0.000 description 1
- JRDYDYXZKFNNRQ-XPUUQOCRSA-N Gly-Ala-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)CN JRDYDYXZKFNNRQ-XPUUQOCRSA-N 0.000 description 1
- KRRMJKMGWWXWDW-STQMWFEESA-N Gly-Arg-Phe Chemical compound NC(=N)NCCC[C@H](NC(=O)CN)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 KRRMJKMGWWXWDW-STQMWFEESA-N 0.000 description 1
- CIMULJZTTOBOPN-WHFBIAKZSA-N Gly-Asn-Asn Chemical compound NCC(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O CIMULJZTTOBOPN-WHFBIAKZSA-N 0.000 description 1
- PABFFPWEJMEVEC-JGVFFNPUSA-N Gly-Gln-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCC(=O)N)NC(=O)CN)C(=O)O PABFFPWEJMEVEC-JGVFFNPUSA-N 0.000 description 1
- PDAWDNVHMUKWJR-ZETCQYMHSA-N Gly-Gly-His Chemical compound NCC(=O)NCC(=O)N[C@H](C(O)=O)CC1=CNC=N1 PDAWDNVHMUKWJR-ZETCQYMHSA-N 0.000 description 1
- FKYQEVBRZSFAMJ-QWRGUYRKSA-N Gly-Ser-Tyr Chemical compound NCC(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 FKYQEVBRZSFAMJ-QWRGUYRKSA-N 0.000 description 1
- XHVONGZZVUUORG-WEDXCCLWSA-N Gly-Thr-Lys Chemical compound NCC(=O)N[C@@H]([C@H](O)C)C(=O)N[C@H](C(O)=O)CCCCN XHVONGZZVUUORG-WEDXCCLWSA-N 0.000 description 1
- SYOJVRNQCXYEOV-XVKPBYJWSA-N Gly-Val-Glu Chemical compound [H]NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O SYOJVRNQCXYEOV-XVKPBYJWSA-N 0.000 description 1
- 206010019233 Headaches Diseases 0.000 description 1
- 208000000616 Hemoptysis Diseases 0.000 description 1
- 244000309467 Human Coronavirus Species 0.000 description 1
- 208000006083 Hypokinesia Diseases 0.000 description 1
- RQJUKVXWAKJDBW-SVSWQMSJSA-N Ile-Ser-Thr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)O)N RQJUKVXWAKJDBW-SVSWQMSJSA-N 0.000 description 1
- 108010065920 Insulin Lispro Proteins 0.000 description 1
- 102100034349 Integrase Human genes 0.000 description 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- LZDNBBYBDGBADK-UHFFFAOYSA-N L-valyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)C(N)C(C)C)C(O)=O)=CNC2=C1 LZDNBBYBDGBADK-UHFFFAOYSA-N 0.000 description 1
- NRFGTHFONZYFNY-MGHWNKPDSA-N Leu-Ile-Tyr Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 NRFGTHFONZYFNY-MGHWNKPDSA-N 0.000 description 1
- RGUXWMDNCPMQFB-YUMQZZPRSA-N Leu-Ser-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O RGUXWMDNCPMQFB-YUMQZZPRSA-N 0.000 description 1
- KLSUAWUZBMAZCL-RHYQMDGZSA-N Leu-Thr-Pro Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N1CCC[C@H]1C(O)=O KLSUAWUZBMAZCL-RHYQMDGZSA-N 0.000 description 1
- TUIOUEWKFFVNLH-DCAQKATOSA-N Leu-Val-Cys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CS)C(O)=O TUIOUEWKFFVNLH-DCAQKATOSA-N 0.000 description 1
- 102000003960 Ligases Human genes 0.000 description 1
- 108090000364 Ligases Proteins 0.000 description 1
- 239000000232 Lipid Bilayer Substances 0.000 description 1
- KNKHAVVBVXKOGX-JXUBOQSCSA-N Lys-Ala-Thr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O KNKHAVVBVXKOGX-JXUBOQSCSA-N 0.000 description 1
- MEQLGHAMAUPOSJ-DCAQKATOSA-N Lys-Ser-Val Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O MEQLGHAMAUPOSJ-DCAQKATOSA-N 0.000 description 1
- SUZVLFWOCKHWET-CQDKDKBSSA-N Lys-Tyr-Ala Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](C)C(O)=O SUZVLFWOCKHWET-CQDKDKBSSA-N 0.000 description 1
- 101710145006 Lysis protein Proteins 0.000 description 1
- 101710085938 Matrix protein Proteins 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 101710127721 Membrane protein Proteins 0.000 description 1
- AFVOKRHYSSFPHC-STECZYCISA-N Met-Ile-Tyr Chemical compound CSCC[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 AFVOKRHYSSFPHC-STECZYCISA-N 0.000 description 1
- 206010027417 Metabolic acidosis Diseases 0.000 description 1
- 101100193635 Mus musculus Rag2 gene Proteins 0.000 description 1
- WUGMRIBZSVSJNP-UHFFFAOYSA-N N-L-alanyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)C(N)C)C(O)=O)=CNC2=C1 WUGMRIBZSVSJNP-UHFFFAOYSA-N 0.000 description 1
- 108010079364 N-glycylalanine Proteins 0.000 description 1
- 108010066427 N-valyltryptophan Proteins 0.000 description 1
- 206010028748 Nasal obstruction Diseases 0.000 description 1
- 108090001074 Nucleocapsid Proteins Proteins 0.000 description 1
- NAXPHWZXEXNDIW-JTQLQIEISA-N Phe-Gly-Gly Chemical compound OC(=O)CNC(=O)CNC(=O)[C@@H](N)CC1=CC=CC=C1 NAXPHWZXEXNDIW-JTQLQIEISA-N 0.000 description 1
- SJRQWEDYTKYHHL-SLFFLAALSA-N Phe-Tyr-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CC3=CC=CC=C3)N)C(=O)O SJRQWEDYTKYHHL-SLFFLAALSA-N 0.000 description 1
- 206010035664 Pneumonia Diseases 0.000 description 1
- PULPZRAHVFBVTO-DCAQKATOSA-N Pro-Glu-Arg Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O PULPZRAHVFBVTO-DCAQKATOSA-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
- SXJOPONICMGFCR-DCAQKATOSA-N Pro-Ser-Lys Chemical compound C1C[C@H](NC1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)O SXJOPONICMGFCR-DCAQKATOSA-N 0.000 description 1
- PRKWBYCXBBSLSK-GUBZILKMSA-N Pro-Ser-Val Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O PRKWBYCXBBSLSK-GUBZILKMSA-N 0.000 description 1
- 206010036790 Productive cough Diseases 0.000 description 1
- 229940096437 Protein S Drugs 0.000 description 1
- 101710188315 Protein X Proteins 0.000 description 1
- 241001112090 Pseudovirus Species 0.000 description 1
- 102000001183 RAG-1 Human genes 0.000 description 1
- 108060006897 RAG1 Proteins 0.000 description 1
- 108091034057 RNA (poly(A)) Proteins 0.000 description 1
- 239000013614 RNA sample Substances 0.000 description 1
- 102000018120 Recombinases Human genes 0.000 description 1
- 108010091086 Recombinases Proteins 0.000 description 1
- 208000036071 Rhinorrhea Diseases 0.000 description 1
- 206010039101 Rhinorrhoea Diseases 0.000 description 1
- 241001678561 Sarbecovirus Species 0.000 description 1
- 206010040070 Septic Shock Diseases 0.000 description 1
- BNFVPSRLHHPQKS-WHFBIAKZSA-N Ser-Asp-Gly Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(O)=O BNFVPSRLHHPQKS-WHFBIAKZSA-N 0.000 description 1
- SFZKGGOGCNQPJY-CIUDSAMLSA-N Ser-Asp-His Chemical compound C1=C(NC=N1)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](CO)N SFZKGGOGCNQPJY-CIUDSAMLSA-N 0.000 description 1
- OLIJLNWFEQEFDM-SRVKXCTJSA-N Ser-Asp-Phe Chemical compound OC[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 OLIJLNWFEQEFDM-SRVKXCTJSA-N 0.000 description 1
- GZFAWAQTEYDKII-YUMQZZPRSA-N Ser-Gly-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CO GZFAWAQTEYDKII-YUMQZZPRSA-N 0.000 description 1
- XXXAXOWMBOKTRN-XPUUQOCRSA-N Ser-Gly-Val Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O XXXAXOWMBOKTRN-XPUUQOCRSA-N 0.000 description 1
- QBUWQRKEHJXTOP-DCAQKATOSA-N Ser-His-Arg Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O QBUWQRKEHJXTOP-DCAQKATOSA-N 0.000 description 1
- ZOPISOXXPQNOCO-SVSWQMSJSA-N Ser-Ile-Thr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H]([C@@H](C)O)C(=O)O)NC(=O)[C@H](CO)N ZOPISOXXPQNOCO-SVSWQMSJSA-N 0.000 description 1
- FUMGHWDRRFCKEP-CIUDSAMLSA-N Ser-Leu-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(O)=O FUMGHWDRRFCKEP-CIUDSAMLSA-N 0.000 description 1
- MUJQWSAWLLRJCE-KATARQTJSA-N Ser-Leu-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O MUJQWSAWLLRJCE-KATARQTJSA-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
- WLJPJRGQRNCIQS-ZLUOBGJFSA-N Ser-Ser-Asn Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O WLJPJRGQRNCIQS-ZLUOBGJFSA-N 0.000 description 1
- PPCZVWHJWJFTFN-ZLUOBGJFSA-N Ser-Ser-Asp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(O)=O)C(O)=O PPCZVWHJWJFTFN-ZLUOBGJFSA-N 0.000 description 1
- GYDFRTRSSXOZCR-ACZMJKKPSA-N Ser-Ser-Glu Chemical compound OC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCC(O)=O GYDFRTRSSXOZCR-ACZMJKKPSA-N 0.000 description 1
- DYEGLQRVMBWQLD-IXOXFDKPSA-N Ser-Thr-Phe Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O)NC(=O)[C@H](CO)N)O DYEGLQRVMBWQLD-IXOXFDKPSA-N 0.000 description 1
- RTXKJFWHEBTABY-IHPCNDPISA-N Ser-Trp-Tyr Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)N[C@@H](CC3=CC=C(C=C3)O)C(=O)O)NC(=O)[C@H](CO)N RTXKJFWHEBTABY-IHPCNDPISA-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
- HAYADTTXNZFUDM-IHRRRGAJSA-N Ser-Tyr-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](C(C)C)C(O)=O HAYADTTXNZFUDM-IHRRRGAJSA-N 0.000 description 1
- HSWXBJCBYSWBPT-GUBZILKMSA-N Ser-Val-Val Chemical compound CC(C)[C@H](NC(=O)[C@@H](NC(=O)[C@@H](N)CO)C(C)C)C(O)=O HSWXBJCBYSWBPT-GUBZILKMSA-N 0.000 description 1
- 101710198474 Spike protein Proteins 0.000 description 1
- 206010051379 Systemic Inflammatory Response Syndrome Diseases 0.000 description 1
- MQCPGOZXFSYJPS-KZVJFYERSA-N Thr-Ala-Arg Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O MQCPGOZXFSYJPS-KZVJFYERSA-N 0.000 description 1
- DIPIPFHFLPTCLK-LOKLDPHHSA-N Thr-Gln-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N1CCC[C@@H]1C(=O)O)N)O DIPIPFHFLPTCLK-LOKLDPHHSA-N 0.000 description 1
- VOHWDZNIESHTFW-XKBZYTNZSA-N Thr-Glu-Cys Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CS)C(=O)O)N)O VOHWDZNIESHTFW-XKBZYTNZSA-N 0.000 description 1
- QQWNRERCGGZOKG-WEDXCCLWSA-N Thr-Gly-Leu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)NCC(=O)N[C@@H](CC(C)C)C(O)=O QQWNRERCGGZOKG-WEDXCCLWSA-N 0.000 description 1
- WPSDXXQRIVKBAY-NKIYYHGXSA-N Thr-His-Glu Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N)O WPSDXXQRIVKBAY-NKIYYHGXSA-N 0.000 description 1
- PRNGXSILMXSWQQ-OEAJRASXSA-N Thr-Leu-Phe Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O PRNGXSILMXSWQQ-OEAJRASXSA-N 0.000 description 1
- VRUFCJZQDACGLH-UVOCVTCTSA-N Thr-Leu-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O VRUFCJZQDACGLH-UVOCVTCTSA-N 0.000 description 1
- FWTFAZKJORVTIR-VZFHVOOUSA-N Thr-Ser-Ala Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(O)=O FWTFAZKJORVTIR-VZFHVOOUSA-N 0.000 description 1
- COYHRQWNJDJCNA-NUJDXYNKSA-N Thr-Thr-Thr Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O COYHRQWNJDJCNA-NUJDXYNKSA-N 0.000 description 1
- OGOYMQWIWHGTGH-KZVJFYERSA-N Thr-Val-Ala Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C)C(O)=O OGOYMQWIWHGTGH-KZVJFYERSA-N 0.000 description 1
- KPMIQCXJDVKWKO-IFFSRLJSSA-N Thr-Val-Glu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O KPMIQCXJDVKWKO-IFFSRLJSSA-N 0.000 description 1
- BKVICMPZWRNWOC-RHYQMDGZSA-N Thr-Val-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)[C@@H](C)O BKVICMPZWRNWOC-RHYQMDGZSA-N 0.000 description 1
- LTLBNCDNXQCOLB-UBHSHLNASA-N Trp-Asp-Ser Chemical compound C1=CC=C2C(C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O)=CNC2=C1 LTLBNCDNXQCOLB-UBHSHLNASA-N 0.000 description 1
- NLLARHRWSFNEMH-NUTKFTJISA-N Trp-Lys-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)N NLLARHRWSFNEMH-NUTKFTJISA-N 0.000 description 1
- ULHASJWZGUEUNN-XIRDDKMYSA-N Trp-Lys-Ser Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(O)=O ULHASJWZGUEUNN-XIRDDKMYSA-N 0.000 description 1
- UABYBEBXFFNCIR-YDHLFZDLSA-N Tyr-Asp-Val Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O UABYBEBXFFNCIR-YDHLFZDLSA-N 0.000 description 1
- NSGZILIDHCIZAM-KKUMJFAQSA-N Tyr-Leu-Ser Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)N NSGZILIDHCIZAM-KKUMJFAQSA-N 0.000 description 1
- ZLFHAAGHGQBQQN-AEJSXWLSSA-N Val-Ala-Pro Chemical compound C[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](C(C)C)N ZLFHAAGHGQBQQN-AEJSXWLSSA-N 0.000 description 1
- BZWUSZGQOILYEU-STECZYCISA-N Val-Ile-Tyr Chemical compound CC(C)[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 BZWUSZGQOILYEU-STECZYCISA-N 0.000 description 1
- RWOGENDAOGMHLX-DCAQKATOSA-N Val-Lys-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C(C)C)N RWOGENDAOGMHLX-DCAQKATOSA-N 0.000 description 1
- UGFMVXRXULGLNO-XPUUQOCRSA-N Val-Ser-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O UGFMVXRXULGLNO-XPUUQOCRSA-N 0.000 description 1
- NZYNRRGJJVSSTJ-GUBZILKMSA-N Val-Ser-Val Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O NZYNRRGJJVSSTJ-GUBZILKMSA-N 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 230000005856 abnormality Effects 0.000 description 1
- 108010081404 acein-2 Proteins 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 208000037892 acute myocardial injury Diseases 0.000 description 1
- 108010024078 alanyl-glycyl-serine Proteins 0.000 description 1
- 108010041407 alanylaspartic acid Proteins 0.000 description 1
- 108010047495 alanylglycine Proteins 0.000 description 1
- 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 1
- 230000004075 alteration Effects 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 230000003915 cell function Effects 0.000 description 1
- 230000036755 cellular response Effects 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 108010016616 cysteinylglycine Proteins 0.000 description 1
- 210000000172 cytosol Anatomy 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- 230000004069 differentiation Effects 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- FSXRLASFHBWESK-UHFFFAOYSA-N dipeptide phenylalanyl-tyrosine Natural products C=1C=C(O)C=CC=1CC(C(O)=O)NC(=O)C(N)CC1=CC=CC=C1 FSXRLASFHBWESK-UHFFFAOYSA-N 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 208000017574 dry cough Diseases 0.000 description 1
- 241001493065 dsRNA viruses Species 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 102000013165 exonuclease Human genes 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 238000012252 genetic analysis Methods 0.000 description 1
- 230000002068 genetic effect Effects 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
- 108010089804 glycyl-threonine Proteins 0.000 description 1
- 108010050848 glycylleucine Proteins 0.000 description 1
- 108010087823 glycyltyrosine Proteins 0.000 description 1
- 231100000869 headache Toxicity 0.000 description 1
- 230000002008 hemorrhagic effect Effects 0.000 description 1
- 239000000833 heterodimer Substances 0.000 description 1
- 238000012165 high-throughput sequencing Methods 0.000 description 1
- 108010085325 histidylproline Proteins 0.000 description 1
- 230000028996 humoral immune response Effects 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 108010091871 leucylmethionine Proteins 0.000 description 1
- 108010057821 leucylproline Proteins 0.000 description 1
- 208000027905 limb weakness Diseases 0.000 description 1
- 231100000861 limb weakness Toxicity 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 238000007481 next generation sequencing Methods 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 108010084572 phenylalanyl-valine Proteins 0.000 description 1
- 210000001236 prokaryotic cell Anatomy 0.000 description 1
- 108010070643 prolylglutamic acid Proteins 0.000 description 1
- 239000012264 purified product Substances 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 210000002345 respiratory system Anatomy 0.000 description 1
- 230000036303 septic shock Effects 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 108010048818 seryl-histidine Proteins 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 238000007671 third-generation sequencing Methods 0.000 description 1
- 102000035160 transmembrane proteins Human genes 0.000 description 1
- 108091005703 transmembrane proteins Proteins 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 108010073969 valyllysine Proteins 0.000 description 1
- 230000007733 viral latency Effects 0.000 description 1
- 210000002845 virion Anatomy 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/08—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses
- C07K16/10—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses from RNA viruses
-
- 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
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
-
- C—CHEMISTRY; METALLURGY
- C40—COMBINATORIAL TECHNOLOGY
- C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
- C40B40/00—Libraries per se, e.g. arrays, mixtures
- C40B40/04—Libraries containing only organic compounds
- C40B40/06—Libraries containing nucleotides or polynucleotides, or derivatives thereof
- C40B40/08—Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/60—Immunoglobulins specific features characterized by non-natural combinations of immunoglobulin fragments
- C07K2317/62—Immunoglobulins specific features characterized by non-natural combinations of immunoglobulin fragments comprising only variable region components
- C07K2317/622—Single chain antibody (scFv)
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
- C07K2317/76—Antagonist effect on antigen, e.g. neutralization or inhibition of binding
Abstract
本发明涉及生物医药技术领域,具体公开了全人源的新冠IgL单链抗体及其应用。本发明通过BCR全长测序,通过分析不同康复者共有而在正常人群中没有的BCR序列,筛选出一类新冠特异性的IgL BCR抗体全长。同时基于抗体全长序列设计特定抗体特异性引物,进行扩增富集后并进行单克隆的制备与后续的克隆筛选测序,获得了BCR序列全长,该序列进行表达后可获得具有中和活性的病毒特异性抗体。通过获得的全人源抗体可以寻找一种能与新冠特异性结合的单链抗体,这类特异识别新冠的单链抗体本身或者其可变区序列,经过直接表达或经基因工程改造成其它的抗体形式,可以作为抗新冠药物开发、疫苗生产、检测标志物开发等新冠相关应用中。
Description
技术领域
本发明涉及生物医药技术领域,具体涉及全人源的新冠IgL单链抗体及其应用。
背景技术
新型冠状病毒是一种具有包膜的、不分节段的正链单股RNA病毒,颗粒呈圆形或椭圆形,直径约80~120nm,属于网巢病毒目冠状病毒科Betacoronavirus。病毒粒子被宿主细胞提供的脂质双层所包裹,其中含有核酸及核衣壳蛋白,有三种主要蛋白:包膜蛋白(E蛋白)、膜蛋白(M蛋白)和刺突蛋白(S蛋白)。此病毒每组基因组长度约三万个核苷酸左右,基因序列显示SARS-CoV-2属于乙型冠状病毒属谱系β(Betacoronavirus Lineageβ,Sarbecovirus)进化树中分支较长的一种病毒,与中华菊头蝠中发现的冠状病毒相似,例如MERS-CoV或SARS-CoV。对病毒的生物遗传学分析显示,同属人类冠状病毒的SARS病毒分离株AY274119相比MERS病毒分离株KC164505、JX869059等在亲缘关系上距离SARS-CoV-2病毒更近。
2019年末,该病爆发并向全球扩散。病毒潜伏期平均大约3-7天,最长不超过14天。大多数患者的表现以下呼吸道症状为主,常见临床表现包括发热、四肢乏力、干咳等症状,其他表现包含鼻塞、流鼻涕、头痛、咽痛、咳血,咳痰、或腹泻等。有部分患者仅表现为低热、轻微乏力等,无肺炎表现。还有部分患者无任何临床表现。病毒感染严重后,会引发多种并发症包含急性呼吸窘迫综合征(ARDS)、脓毒症休克、全身炎症反应综合征、难以纠正的代谢性酸中毒、急性心肌损伤,和出凝血功能障碍等。
针对这场危及全人类的群体大流行,揭示人体对新冠侵染后的免疫过程,获取新冠病毒的B细胞受体对于新冠特异性药物的筛选、病毒疫苗的生产研发及其重要。B细胞受体(B cell receptor,BCR)是B细胞抗原识别决定性表面分子,其本质是一种膜表面免疫球蛋白(membrane immunoglobulin,mIg)。BCR具有抗原结合特异性,每个个体的BCR多样性高达5×10^13,构成容量巨大的BCR库,赋予个体识别各种抗原、产生特异性抗体的巨大潜能。
BCR的结构包括重链和轻链。BCR的重链(heavy chain,H)由65~100种可变区(VH)、2种多变区(DH)、6种结合区(JH)和恒定区(CH)四部分基因片段组成;轻链(lightchain,L)由可变区、结合区和恒定区三部分基因片段组成。发育过程中的B细胞在重组酶(RAG1、RAG2)作用下,形成了多样性高达1~2×10^11的BCR。同时,由其形成互补决定区(complementarities determining region,CDR):CDR1、CDR2和CDR3区氨基酸序列的多样性,特别是编码CDR3的基因,由于其位于轻链V、J或重链V、D、J片段的连接处,可以通过V(D)J的重排和(或)两个基因片段的连接间丢失或插入数个核苷酸,进一步增加BCR的多样性,而形成具有功能的BCR编码基因(B细胞克隆)。
BCR测序是通过高通量测序技术检测靶向扩增后的BCR重链和轻链,全面解析BCR基因重排碱基序列,以及各序列的丰度的测序技术。BCR测序常用于评价各种免疫相关疾病和遗传性突变引起的某个物种所有B细胞或特定B细胞激活介导的细胞免疫反应中BCR基因重排碱基序列,以及各序列的丰度,用于研究不同B细胞克隆的转录情况和相互间关系,从而揭示更深层次的B细胞功能特异性,继而解释体液免疫应答耐受以及高频突变在B细胞应答识别抗原异常等相关生命现象。传统的BCR测序都是采用使用Illumina的测序仪进行2×300bp或2×150bp的双端测序方法来对BCR进行测序,该方法的测序准确率高,但对于部分长度超过600bp的BCR序列,该方法仅能获得两端的序列,会存在中间部分关键可变区序列缺失的问题。
PacBio公司正式推出Sequel II新一代测序系统后,依赖于PacBio SMRT测序技术独有的CCS测序模式,能够通过滚环测序提升reads准确度,同时结合聚合酶试剂的优化,酶读长大幅提升,在获得高精准度HiFi reads的同时能够保证10kb以上的插入片段读长,将解决原有的Illumina等二代测序平台下无法完整覆盖整个片段区域的问题。通过进行HiFi测序,可以获得准确性达到99.5%以上的长读长序列。
目前,针对新冠病毒侵染人体后引起的BCR免疫反应,常规的方法是采用二代测序平台对抗体的可变区进行测序后筛选新冠相关抗体,但是受限于测序的读长,仅能够获得可变区的部分序列,无法进行准确的抗体类型识别与编码翻译识别,而基于三代测序平台PacBio的新冠病毒BCR全长扩增测序可轻松读取病毒侵染后的BCR全长序列,突破了二代测序读长较短的局限性,提高了针对新冠病毒特异性BCR抗体的分辨能力,极大的提高了新冠特异性BCR抗体鉴定的分辨率与准确率。特别是在PacBio公司正式推出Sequel II新一代测序系统后,依赖于PacBio SMRT测序技术独有的HiFi测序模式,能够通过滚环测序提升reads准确度,目前采用HiFi技术进行BCR抗体全长测序的准确率可以达到99.5%以上,通过进行HiFi测序,可以获得极高准确性的新冠BCR抗体全长序列,用于新冠相关BCR抗体序列筛选、基于全长抗体序列的表达翻译与后续新冠中和反应评估等。
BCR一般包括与膜结合的免疫球蛋白分子和Ig-α/Ig-β信号转导组件,通过二硫键连接着免疫球蛋白分子和信号转导组件。BCR包括以下两部分:1.某一亚型(IgD、IgM、IgL、IgG或者IgE)的膜结合免疫球蛋白(mIg)。除了C端疏水的膜结合区和胞内区,这些膜结合免疫球蛋白和分泌型的免疫球蛋白单体是相同的,有两条重链(IgHs)和两条轻链(IgLs);2.信号转导组件:Ig-α/Ig-β的异二聚体(CD79),由二硫键连接。两个亚基都是跨膜蛋白,在胞内区都有免疫受体酪氨酸的活化基序(ITAM)。
免疫球蛋白有两个轻链,称为lambda型或kappa型,还有两条重链,它们的结构中有特定的变量,以适应体内的结合和精确的功能。轻链之所以“轻”,是因为它们由大约220个氨基酸组成,而重链通常由500多个组成。完整的免疫球蛋白在人体系统中承担着两个不同的角色;免疫球蛋白可作为细胞表面的抗原受体,或在细胞液中自由循环,以识别和消除抗原化合物或病原体。免疫球蛋白的功能决定与其发生和分化过程有关。免疫球蛋白是由作为抗体的白细胞。一种抗体分子是由一种称为B淋巴细胞(B细胞)的白细胞产生的。每个产生免疫球蛋白的B细胞只被编码来表达和产生一种单一形式的免疫球蛋白轻链。卡帕链在2号染色体上有一个由免疫球蛋白编码的特定位点kappa基因座(IgK)和另一种基因lambda链,在22号染色体上有一个由免疫球蛋白lambda位点(IgL)编码的位点。血清免疫球蛋白轻链中,kappa链与lambda链的比率约为2:1。
目前为止,未能通过筛选方式获得新冠特异性的IgL BCR抗体全长。
发明内容
本发明目的在于提供一种全人源的新冠IgL单链抗体,这类特异识别新冠的单链抗体本身或者其可变区序列,经过直接表达或经基因工程改造成其它的抗体形式后,可以作为抗新冠药物开发、疫苗生产、检测标志物开发等进行新冠相关应用。
本发明提供一种全人源的新冠IgL单链抗体,包括如SEQ ID No:4-SEQ ID No:6中至少一种所示的氨基酸序列。
本发明还提供,编码上述新冠IgL单链抗体的基因序列,优选地,包括如SEQ IDNo:1-SEQ ID No:3中至少一种与上述氨基酸对应的核苷酸序列。
本发明还提供,包含上述新冠IgL单链抗体中所述基因序列的文库。
本发明还提供,上述文库的制备方法,对不同新冠康复者共有而在正常人群中没有的BCR序列进行分析并筛选出新冠特异性的IgL BCR抗体序列。
进一步地,所述分析筛选过程为:对新冠康复者与正常人群进行BCR全长测序,将测序的原始数据进行HiFi的一致性校正后,获得了质量值在Q20以上的BCR全长一致性序列,经过与BCR数据库中的抗体恒定区序列进行比对后,获得了每个样本测序数据中的不同类别的BCR抗体序列。
本发明还提供,包含上述新冠IgL单链抗体中所述基因序列的表达载体。
本发明还提供,包含上述新冠IgL单链抗体中所述基因序列的宿主细胞。
本发明还提供上述新冠IgL单链抗体在制备新冠病毒治疗药物、药物载体及检测标记物中的应用。
相比现有技术,本发明的有益效果为:
1.本发明通过使用BCR全长扩增子测序的方法对新冠康复者与健康人群进行了BCR全长扩增建库测序,获得了质量值在Q20以上的BCR全长扩增子序列,获得的BCR序列包括完整的启动子至终止密码子的的区域,所获得的序列为全人源,无需进一步进行整合即可进行后续的表达验证。
2.本发明通过PacBio HiFi测序获得高质量的BCR序列,经与数据库进行比对后,根据人的BCR序列在恒定区的保守性,对获得的BCR全长序列进行转录方向校正与分类,构建新冠康复者与建库人群不同类别的BCR抗体全长数据库;传统的二代BCR测序技术仅进行部分可变区的测序,无法准确进行抗体的聚类筛选。
3.本发明针对获取的不同类型的BCR序列,基于不同类别抗体恒定区BCR序列的一致性,通过定位位于恒定区上的终止密码子位置,将获得的抗体直接翻译为蛋白质氨基酸序列,在蛋白质水平上对抗体进行比较;传统的二代BCR测序方法不涉及BCR序列的恒定区,无法进行准确的翻译,仅能在DNA序列的水平上进行比较。
4.本发明通过对新冠康复者与正常人群的不同类别的抗体蛋白质氨基酸序列进行比较,找到新冠康复者共有而在正常人群中没有的BCR抗体,这些抗体为新冠特异性抗体;其抗体DNA序列为全人源的新冠特异性抗体全长序列,在经过直接表达或经基因工程改造成其它的抗体形式后,可以作为抗新冠药物开发、疫苗生产、检测标志物开发等进行新冠相关应用中;传统的新冠相关应用均是基于可变区的部分序列进行基因工程改造后再进行表达,由于其序列并非全人源,且必须经过改造,可能引入某些未知的安全问题。
5.本发明通过对获取的新冠特异性抗体序列设计特异性引物,该引物扩增区域包括启动子至终止密码子的全部区域,使用该引物在新冠康复者的cDNA作为模板进行PCR扩增后,将扩增产物转移至大肠杆菌表达载体中,通过构建单克隆后进行一代测序,基于扩增片段大小筛选获取的目标序列进行测序,在测序数据中找到了单一的新冠特异性的抗体DNA序列,获取的DNA序列为单一的纯的全人源特定抗体DNA序列,无需进行人工合成即可直接作为抗新冠药物开发、疫苗生产、检测标志物开发的原始DNA反应物。
附图说明
为了更清楚地说明本发明实施例的技术方案,下面将对实施例中所需要使用的附图作简单地介绍,应当理解,以下附图仅示出了本发明的某些实施例,因此不应被看作是对范围的限定,对于本领域普通技术人员来讲,在不付出创造性劳动的前提下,还可以根据这些附图获得其他相关的附图。
图1是本发明新冠相关特异性抗体筛选与获取原理示意图。
图2是本发明特异性引物扩增富集新冠相关BCR序列电泳图示意图。
图3是本发明单克隆制备培养示意图。
图4是本发明单克隆扩增筛选电泳图。
具体实施方式
以下实例用于说明本发明,但不限制本发明的范围。在不背离本发明精神和实质的前提下,对本发明的方法、步骤或条件所作的修改或替换,均属于本发明的范围。
如图1所示,本发明提供的全人源的新冠单链抗体获取流程如下:
1.通过进行新冠康复者与健康人群的BCR全长扩增子建库测序,获取质量值Q20以上的BCR全长序列,分别构建新冠康复者与健康人群的BCR抗体库;
2..在获得高质量的BCR抗体序列后,与抗体数据库进行比对,构建BCR抗体恒定区序列确定抗体类别,由于同类型的BCR抗体相似性非常高,且进行翻译时翻译的终止密码子位置也是一致,基于终止密码子位置将BCR抗体序列翻译为蛋白质氨基酸序列;
3.通过比较新冠康复者共有,而健康人群没有的BCR蛋白质氨基酸序列,找到新冠特异性相关的不同类别的BCR序列;
4.基于找到的新冠特异性序列设计特异引物,进行扩增特异性富集新冠相关的抗体序列;
5.将抗体序列转移至感受态细胞中,制备单克隆,挑选单克隆进行扩增后,选择特定片段大小的扩增产物进行一代测序,基于一代测序的结果筛选获得的全人源的新冠相关的BCR序列。
在本发明中,获得了新冠特异性IgL序列1条,获得的新冠特异性IgL序列如SEQ IDNo:1-3所示,经翻译后的氨基酸序列如SEQ ID No:4-6所示。所有的序列均包含VDJ区域,在5’端包括起始密码子ATG,在3’端包括终止密码子TGA,该抗体序列为全人源序列,可安全的应用于后续疫苗生产与抗体药物研发等应用中。
根据本发明的新冠特异性IgL中特异性核苷酸或氨基酸序列,可在体外人工合成与此相同的抗体轻重链基因的核苷酸序列或编码相同氨基酸的核苷酸序列,从而获得相同的抗体基因或用于相关基因的改造,而获得IgL抗体或相关蛋白。
在本发明中,提出的包含上述新冠IgL单链抗体中所述基因序列的文库,所述文库为:通过对新冠康复者与健康人群的BCR全长扩增子建库测序,获取质量值Q20以上的BCR全长序列,分别构建新冠康复者与健康人群的BCR抗体库;经与数据库进行比对后,根据人的BCR序列在恒定区的保守性,对获得的BCR全长序列进行转录方向校正与分类,构建新冠康复者与建库人群不同类别的BCR抗体全长数据库。
在本发明中,提出包含所述基因序列的表达载体,根据本领域的常识,载体可以为原核细胞表达载体、真核细胞表达载体或昆虫细胞表达载体。
在本发明中,提出包含所述表达载体的宿主细胞,根据本领域的常识,所述的宿主细胞可以为原核表达细胞、真核表达细胞或昆虫细胞,所述原核表达细胞优选大肠杆菌。
实施例一:新冠康复者与正常人群BCR全长扩增测序
新冠康复者与正常人群的BCR全长扩增测序参照前期申请的专利方法(公开号CN111662970A)进行,主要流程如下:
1.全血样本Total RNA的提取
取新鲜全血样本1mL,使用TriZol LS进行全血样本Total RNA提取,提取后使用Nanidrop 2000C测定RNA样本的浓度与纯度,使用Agilent 2100测定样本的完整性,达到合格标准的样本(总量>1μg,完整性RIN值>7)的样本进行后续实验。
2.总RNA中cDNA第一链的合成
实验操作流程如下:
1)Oligo dT逆转录引物与poly(A)结合,如表1所示。
表1:
轻弹混匀,瞬时离心,70℃孵育5min后立即置于冰上。
2)逆转录合成cDNA第一链,配制表2中的反应。
表2:
轻弹混匀,瞬时离心,42℃孵育75min。反应完成后立即置于冰上,加入1μL的BCRTemplate Switching Oligo,轻弹混匀,瞬时离心,42℃孵育15min。
3.BCR cDNA的全长扩增
BCR cDNA的全长扩增包括两轮半巢式扩增反应,第一轮扩增进行BCR序列的初步富集,第二轮扩增时采用内部的巢式引物,进一步提高扩增的特异性,使扩增条带单一。
1)BCR cDNA全长的第一轮PCR扩增
取一新的0.2mL PCR管,加入表3中的试剂。
表3:
充分混匀,瞬时离心,置于PCR仪上进行PCR反应:98℃2min;98℃20s、65℃15s、72℃45s,18cycles;72℃5min。
2)BCR cDNA全长的第二轮PCR扩增
取一新的0.2mL PCR管,加入表4中的试剂。
表4:
10组混合引物均需要单独进行扩增,可以提高扩增反应的稳定性。
充分混匀,瞬时离心,置于PCR仪上进行PCR反应:98℃2min;98℃20s、65℃15s、72℃30s,20cycles;72℃5min。
反应结束后按照AMPure磁珠的使用说明对扩增产物进行磁珠纯化,最后使用10μL洗脱缓冲液洗脱,取1μL纯化产物,使用无核酸酶水稀释5倍后进行Qubit定量。
4.BCR全长扩增子片段混样
根据Qubit定量结果,将同一样本的不同扩增产物进行等量混样,要求混样后的总量在1μg以上,用于后续建库。
5.文库构建
1)末端修复
取1μg的全基因组扩增子样本,进行末端修复反应体系的配制,配制表5中的反应。
表5:
充分混匀,瞬时离心,20℃孵育30min。
反应完成后,按照AMPure磁珠的使用说明进行1X磁珠纯化,去除反应时加入的酶与Buffer,最后使用14μL洗脱缓冲液洗脱,获得片段末端加A的粘性末端。
2)连接带barcode的测序接头
通过末端修复与加A后,在加入具有与A末端匹配的带barcode的测序接头,在连接酶的作用下即可实现接头的连接。反应体系如表6。
表6:
混匀,瞬时离心,20℃孵育60min,反应结束后65℃孵育10min,置于冰上。进行外切酶消化,反应体系如表7。
表7:
混匀,瞬时离心,37℃孵育60min,置于冰上。按照AMPure磁珠的使用说明进行磁珠纯化,最后使用20μL洗脱缓冲液洗脱,获得的适用于PacBio测序平台的哑铃型的环状文库。
3)文库质检与上机测序
取1μL文库进行Qubit定量,获得文库浓度;取1μL文库进行安捷伦2100的片段大小分析,将全长扩增文库在PacBio Sequel II测序平台上进行混合测序,每个样本获得了约60G的测序数据。
实施例二:新冠康复者与正常人群BCR免疫组库分析,获得新冠特异性IgL单链抗体
将测序的原始数据进行HiFi的一致性校正后,获得了质量值在Q20以上的BCR全长一致性序列,经过与BCR数据库中的抗体恒定区序列进行比对后,获得了每个样本测序数据中的不同类别的BCR抗体序列,同时基于恒定区的终止密码子位置,将DNA序列翻译为蛋白质多肽序列。通过比较新冠康复者共有的,而正常人群中没有的BCR多肽序列,获得了新冠特异性相关的抗体全长。本次获得的新冠特异性IgL序列3条,所有的序列均包含VDJ区域,在5’端包括起始密码子ATG,在3’端包括终止密码子TGA,该抗体序列为全人源序列,可安全的应用于后续疫苗生产与抗体药物研发等应用中。获得的新冠特异性IgL序列如SEQ IDNo:1-3所示,经翻译后的氨基酸序列如SEQ ID No:4-6所示。
实施例三:基于以新冠康复者样本设计特异性引物,进行单克隆BCR抗体库的构建
基于抗体库筛选的IgL抗体序列,在起始密码子的与终止密码子的外测设计特异性引物,通过PCR扩增特异性富集目标抗体序列。由于抗体存在多重重组的特性,获取的扩增产物是一类与引物匹配的抗体序列集合,无法直接进行一代测序与后续应用,需要将抗体序列进行克隆实验,获取单个抗体序列的单克隆,并挑选单克隆进行测序,验证所获得的单克隆序列为目标抗体。具体流程如下:
1.目标抗体IgL抗体序列引物设计
基于测序筛选的目标序列抗体设计引物,该引物扩增的目标片段包括起始密码子与终止密码子区域,覆盖整个抗体表达区域的全长,具体序列如表8所示。
表8:IgL抗体序列引物
其中,Primer_ID_01~02为R端通用引物,该引物在恒定区内,所有IgL抗体序列均可与该引物进行匹配,使用时混合在一起作为R端通用引物使用;Primer_ID_03~08为F端引物,该引物序列基于筛选的新冠相关特异性抗体设计,与R端引物组合在一起为一对引物,用于目标抗体序列的扩增。
2.扩增富集获取新冠相关特异性抗体
本步骤使用的扩增模板为新冠康复者样本的单链cDNA样本。取一新的0.2mL PCR管,加入如表9所示试剂。
表9:
充分混匀,瞬时离心,置于PCR仪上进行PCR反应:98℃2min;98℃20s、65℃15s、72℃120s,35cycles;72℃5min。
反应结束后进行电泳检测,电泳结果示意图如图2所示。切胶获取片段大小在1.5~2k的目标特异性片段。
3.目标片段的单克隆制备
本步骤使用表10中的试剂。
表10:
LB固体培养基配制:取本品8克,溶解于250ml蒸馏水中,121℃高压灭菌15分钟,至不烫手时加入250μL氨苄青霉素,混匀倒平板备用(每个平板15mL左右)。
LB液体培养基配制:取本品1克,溶解于40ml蒸馏水中,121℃高压灭菌15分钟,分装至2mL无菌离心管中备用。
配制好培养基后进行载体连接反应,配制如表11中的反应体系。
表11:
轻弹管底混匀,低速瞬时离心收集所有液体于离心管底,室温25℃反应5min。反应结束后,将离心管置于冰上。
将Fast-T1感受态细胞从-70℃拿出,迅速置于冰上融化,取20μL感受态细胞加入目的载体连接反应产物,轻弹管壁混匀(避免用枪吸打),冰上静置30min。
42℃水浴热激30s后,迅速置于冰上静置2min,切勿摇动离心管。
向离心管中加入200μL LB液体培养基(不含抗生素),混匀后置于37℃,200rpm摇床中复苏5min。
复苏后颠倒混匀直接取200μL,涂布在含氨苄青霉素的LB固体培养基平板上,将平板正置于37℃培养箱10min,待菌液被完全吸收后,倒置平板,过夜培养,单克隆制备培养结果示意图如图3所示。
4.单克隆筛选与测序鉴定
在平板培养基上挑选单克隆菌落至10μL ddH2O中混匀作为模板;使用2×RapidTaq Master Mix(Vazyme#P222)进行扩增,反应体系如表12所示。
表12:
放置在PCR仪上进行如表13所示的扩增反应程序。
表13:
将扩增获得的扩增产物进行凝胶电泳检测,获得图4所示的电泳图,挑选扩增条带亮度高、扩增条带单一且片段大小在1.5~5k的扩增产物进行双端单克隆一代测序鉴定,根据一代测序的序列与目标抗体序列进行一致性比对,选择序列完全一致的单克隆进一步进行扩增放大,即获得了高纯度的单一的新冠相关抗体特异性序列,该序列可以直接转移至假病毒体系中,进行后续的抗体效价验证,也可以直接转移至抗体表达体系中进行表达。
序列表
<110> 武汉菲沙基因组医学有限公司
<120> 全人源的新冠IgL单链抗体及其应用
<160> 14
<170> SIPOSequenceListing 1.0
<210> 2
<211> 714
<212> DNA
<213> IgL
<400> 2
atggcctggt ctcctctcct cctcactctc ctcgctcact gcacagggtc ctgggcccag 60
tctgtgctga cgcagccgcc ctcagtgtct ggggccccag ggcagagggt caccatctcc 120
tgcactggga ccagctccaa cctcggggca ggttatgatg tacactggta ccagcagctt 180
ccaggaacag cccccaaact cctcatctat ggtaacaaca agcggccctc agggatccct 240
ggccgattct ctggctccaa gtctggcacc tcagcctccc tggccatcac tgggctccag 300
gctgaggatg aggctgatta ttactgccag tcctatgaca gcaacctgag tggttctgtg 360
gtattcggcg gagggaccaa gctgaccgtc ctaggtcagc ccaaggctgc cccctcggtc 420
actctgttcc cgccctcctc tgaggagctt caagccaaca aggccacact ggtgtgtctc 480
ataagtgact tctacccggg agccgtgaca gtggcctgga aggcagatag cagccccgtc 540
aaggcgggag tggagaccac cacaccctcc aaacaaagca acaacaagta cgcggccagc 600
agctatctga gcctgacgcc tgagcagtgg aagtcccaca gaagctacag ctgccaggtc 660
acgcatgaag ggagcaccgt ggagaagaca gtggccccta cagaatgttc atag 714
<210> 2
<211> 702
<212> DNA
<213> IgL
<400> 2
atggcctgga ccgttctcct cctcggcctc ctctctcact gcacaggctc tgtgacctcc 60
tatgtgctga ctcagccacc ctcagtgtca gtggccccag gaaagacggc caggatttcc 120
tgtgggggac acaacattgg aagtaagagt gttcactggt accagcagaa gccaggccag 180
gcccctgtgc tggtcatcta ttctgatggc gaccggccct cagggatccc tgagcgattc 240
tctggctcca actctgcgaa ggcggccacc ttgaccatca gcacggtcga ggccggggat 300
gaggccgact attactgtca ggtgtgggat agtagtagtg atcacgtggt attcggcgga 360
gggaccaagc tgaccgtcct aggtcagccc aaggctgccc cctcggtcac tctgttcccg 420
ccctcctctg aggagcttca agccaacaag gccacactgg tgtgtctcat aagtgacttc 480
tacccgggag ccgtgacagt ggcctggaag gcagatagca gccccgtcaa ggcgggagtg 540
gagaccacca caccctccaa acaaagcaac aacaagtacg cggccagcag ctatctgagc 600
ctgacgcctg agcagtggaa gtcccacaga agctacagct gccaggtcac gcatgaaggg 660
agcaccgtgg agaagacagt ggcccctaca gaatgttcat ag 702
<210> 3
<211> 711
<212> DNA
<213> IgL
<400> 3
atggcctggg ctctgctgct cctcactctc ctcactcagg acacagggtc ctgggcccag 60
tctgccctga ctcagcctgc ctccgtgtct gggtctcctg gacagtcgat caccatctcc 120
tgcactggaa ccagcagtga tgttgggagt tataaccttg tctcctggta ccaacagcac 180
ccaggcaaag cccccaaact catgatttat gagggcagta agcggccctc aggggtttct 240
aatcgcttct ctggctccaa gtctggcaac acggcctccc tgacaatctc tgggctccag 300
gctgaggacg aggctgatta ttactgctgc tcatatgcag gtagtagcac tttcgtggta 360
ttcggcggag ggaccaagct gaccgtccta ggtcagccca aggctgcccc ctcggtcact 420
ctgttcccgc cctcctctga ggagcttcaa gccaacaagg ccacactggt gtgtctcata 480
agtgacttct acccgggagc cgtgacagtg gcctggaagg cagatagcag ccccgtcaag 540
gcgggagtgg agaccaccac accctccaaa caaagcaaca acaagtacgc ggccagcagc 600
tatctgagcc tgacgcctga gcagtggaag tcccacagaa gctacagctg ccaggtcacg 660
catgaaggga gcaccgtgga gaagacagtg gcccctacag aatgttcata g 711
<210> 4
<211> 237
<212> PRT
<213> IgL_ID_001_translation
<400> 4
Met Ala Trp Ser Pro Leu Leu Leu Thr Leu Leu Ala His Cys Thr Gly
1 5 10 15
Ser Trp Ala Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala
20 25 30
Pro Gly Gln Arg Val Thr Ile Ser Cys Thr Gly Thr Ser Ser Asn Leu
35 40 45
Gly Ala Gly Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala
50 55 60
Pro Lys Leu Leu Ile Tyr Gly Asn Asn Lys Arg Pro Ser Gly Ile Pro
65 70 75 80
Gly Arg Phe Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile
85 90 95
Thr Gly Leu Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr
100 105 110
Asp Ser Asn Leu Ser Gly Ser Val Val Phe Gly Gly Gly Thr Lys Leu
115 120 125
Thr Val Leu Gly Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro
130 135 140
Pro Ser Ser Glu Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu
145 150 155 160
Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp
165 170 175
Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln
180 185 190
Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu
195 200 205
Gln Trp Lys Ser His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly
210 215 220
Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser
225 230 235
<210> 5
<211> 233
<212> PRT
<213> IgL_ID_002_translation
<400> 5
Met Ala Trp Thr Val Leu Leu Leu Gly Leu Leu Ser His Cys Thr Gly
1 5 10 15
Ser Val Thr Ser Tyr Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala
20 25 30
Pro Gly Lys Thr Ala Arg Ile Ser Cys Gly Gly His Asn Ile Gly Ser
35 40 45
Lys Ser Val His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu
50 55 60
Val Ile Tyr Ser Asp Gly Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe
65 70 75 80
Ser Gly Ser Asn Ser Ala Lys Ala Ala Thr Leu Thr Ile Ser Thr Val
85 90 95
Glu Ala Gly Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser
100 105 110
Ser Asp His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly
115 120 125
Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu
130 135 140
Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe
145 150 155 160
Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val
165 170 175
Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys
180 185 190
Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser
195 200 205
His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu
210 215 220
Lys Thr Val Ala Pro Thr Glu Cys Ser
225 230
<210> 6
<211> 236
<212> PRT
<213> IgL_ID_003_translation
<400> 6
Met Ala Trp Ala Leu Leu Leu Leu Thr Leu Leu Thr Gln Asp Thr Gly
1 5 10 15
Ser Trp Ala Gln Ser Ala Leu Thr Gln Pro Ala Ser Val Ser Gly Ser
20 25 30
Pro Gly Gln Ser Ile Thr Ile Ser Cys Thr Gly Thr Ser Ser Asp Val
35 40 45
Gly Ser Tyr Asn Leu Val Ser Trp Tyr Gln Gln His Pro Gly Lys Ala
50 55 60
Pro Lys Leu Met Ile Tyr Glu Gly Ser Lys Arg Pro Ser Gly Val Ser
65 70 75 80
Asn Arg Phe Ser Gly Ser Lys Ser Gly Asn Thr Ala Ser Leu Thr Ile
85 90 95
Ser Gly Leu Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Cys Ser Tyr
100 105 110
Ala Gly Ser Ser Thr Phe Val Val Phe Gly Gly Gly Thr Lys Leu Thr
115 120 125
Val Leu Gly Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro
130 135 140
Ser Ser Glu Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile
145 150 155 160
Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser
165 170 175
Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser
180 185 190
Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln
195 200 205
Trp Lys Ser His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser
210 215 220
Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser
225 230 235
<210> 7
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 7
gagggtttat tgagtgcagg 20
<210> 8
<211> 19
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 8
gatcctgcag ctctagtct 19
<210> 9
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 9
aagcagcact ggtggtgcct c 21
<210> 10
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 10
aggcagcact caggacaatc tc 22
<210> 11
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 11
tctcgggacg tctccaccat 20
<210> 12
<211> 23
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 12
atggcctgga ccgttctcct cct 23
<210> 13
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 13
acaatctcca gcatggcctg g 21
<210> 14
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 14
atggcctggg ctctgctgct cctca 25
Claims (9)
1.一种全人源的新冠IgL单链抗体,其特征在于,包括如SEQ ID No:4-SEQ ID No:6中至少一种所示的氨基酸序列。
2.编码权利要求1所述的新冠IgL单链抗体的基因序列。
3.根据权利要求2所述的新冠IgL单链抗体,其特征在于,包括如SEQ ID No:1-SEQ IDNo:3中至少一种与所述氨基酸对应的核苷酸序列。
4.包含权利要求2所述新冠IgL单链抗体中所述基因序列的文库。
5.权利要求4所述文库的制备方法,其特征在于,对不同新冠康复者共有而在正常人群中没有的BCR序列进行分析并筛选出新冠特异性的IgL BCR抗体序列。
6.根据权权利要求5所述的制备方法,其特征在于,所述分析筛选过程为:对新冠康复者与正常人群进行BCR全长测序,将测序的原始数据进行HiFi的一致性校正后,获得了质量值在Q20以上的BCR全长一致性序列,经过与BCR数据库中的抗体恒定区序列进行比对后,获得了每个样本测序数据中的不同类别的BCR抗体序列。
7.包含权利要求3所述新冠IgL单链抗体中所述基因序列的表达载体。
8.包含权利要求3所述新冠IgL单链抗体中所述基因序列的宿主细胞。
9.权利要求1所述的新冠IgL单链抗体在制备新冠病毒治疗药物、药物载体及检测标记物中的应用。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202110545795.4A CN113214389A (zh) | 2021-05-19 | 2021-05-19 | 全人源的新冠IgL单链抗体及其应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202110545795.4A CN113214389A (zh) | 2021-05-19 | 2021-05-19 | 全人源的新冠IgL单链抗体及其应用 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN113214389A true CN113214389A (zh) | 2021-08-06 |
Family
ID=77093149
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202110545795.4A Pending CN113214389A (zh) | 2021-05-19 | 2021-05-19 | 全人源的新冠IgL单链抗体及其应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN113214389A (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11732030B2 (en) | 2020-04-02 | 2023-08-22 | Regeneron Pharmaceuticals, Inc. | Anti-SARS-CoV-2-spike glycoprotein antibodies and antigen-binding fragments |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111732655A (zh) * | 2020-07-01 | 2020-10-02 | 中国人民解放军军事科学院军事医学研究院 | 靶向RBD的高中和活性抗SARS-CoV-2全人源单克隆抗体及应用 |
CN112794899A (zh) * | 2021-03-16 | 2021-05-14 | 易康生物(苏州)有限公司 | 一种抗新型冠状病毒的全人源单克隆中和抗体及其应用 |
-
2021
- 2021-05-19 CN CN202110545795.4A patent/CN113214389A/zh active Pending
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111732655A (zh) * | 2020-07-01 | 2020-10-02 | 中国人民解放军军事科学院军事医学研究院 | 靶向RBD的高中和活性抗SARS-CoV-2全人源单克隆抗体及应用 |
CN112794899A (zh) * | 2021-03-16 | 2021-05-14 | 易康生物(苏州)有限公司 | 一种抗新型冠状病毒的全人源单克隆中和抗体及其应用 |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11732030B2 (en) | 2020-04-02 | 2023-08-22 | Regeneron Pharmaceuticals, Inc. | Anti-SARS-CoV-2-spike glycoprotein antibodies and antigen-binding fragments |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN112010965B (zh) | 一种针对新冠病毒SARS-CoV-2棘突蛋白RBD区的单克隆抗体及其应用 | |
CN111778218B (zh) | 噬菌体展示抗体库及基于其淘筛获得的针对新冠病毒SARS-CoV-2的单克隆抗体 | |
CN113292640B (zh) | 一种产生广谱交叉中和活性的重组新型冠状病毒rbd三聚体蛋白疫苗、其制备方法和应用 | |
CN113336844B (zh) | 一种靶向新冠病毒n蛋白的鲨鱼单域抗体及其制备方法和应用 | |
CN113150136B (zh) | 新型冠状病毒n蛋白单克隆抗体的制备 | |
CN113912710B (zh) | 抗新型冠状病毒n蛋白的单克隆抗体及其应用 | |
CN113234149B (zh) | 全人源的新冠IgA单链抗体及其应用 | |
CN113416710B (zh) | 一种鼠抗mcr-1蛋白杂交瘤细胞株,单克隆抗体及应用 | |
CN112010964A (zh) | 一种新冠病毒羊驼抗体及其制备方法和应用 | |
CN113527522A (zh) | 一种新冠病毒三聚体重组蛋白、DNA、mRNA及应用和mRNA疫苗 | |
WO2023125520A1 (zh) | SARS-CoV-2α、β、γ和δ突变株骆驼源高亲和力纳米抗体 | |
CN114874995B (zh) | 猪瘟病毒2型Erns蛋白的单克隆抗体杂交瘤细胞株及应用 | |
CN114250203B (zh) | 一种鼠抗mcr-1/mcr-2蛋白杂交瘤细胞株,单克隆抗体及应用 | |
CN111944044A (zh) | 一种抗ASFV-p30蛋白的纳米抗体及其制备方法和应用 | |
CN108659124B (zh) | 一种抗猪流行性腹泻病毒的单链抗体及其应用 | |
CN113214389A (zh) | 全人源的新冠IgL单链抗体及其应用 | |
CN112500479B (zh) | 一种犬ⅱ型腺病毒重组蛋白单克隆抗体的制备 | |
CN112608385B (zh) | 一种犬脑钠肽(bnp)单克隆抗体的制备 | |
CN112898421A (zh) | 一种新冠病毒s1-rbd蛋白羊驼纳米抗体及其制备方法和应用 | |
JPS5944399A (ja) | 新規dna | |
CN114213532A (zh) | 高亲和力的抗鸡传染性法氏囊病毒的scFv抗体的制备及其应用 | |
CN110655572A (zh) | 一种抗丝状病毒gp蛋白的单克隆抗体及其应用 | |
CN113105544A (zh) | 全人源的新冠IgG4单链抗体及其应用 | |
CN111704665B (zh) | 一种针对H3N2犬流感病毒的重组犬源化抗体scFv-Fc | |
CN113173989B (zh) | 抗纤维蛋白-2抗体序列、四肽链分子及免疫球蛋白分子 |
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 |