Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>)
Tumor necrosis factor α (TNFα), an important clinical testing factor and drug target, can trigger serious autoimmune diseases and inflammation. Thus, the TNFα antibodies have great potential application in diagnostics and therapy fields. The variable binding domain of IgNAR (VNAR), the shark single...
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| Format: | Article |
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MDPI AG
2022-04-01
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| Series: | Marine Drugs |
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| Online Access: | https://www.mdpi.com/1660-3397/20/5/307 |
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| author | Linfei Zhao Mingliang Chen Xiaona Wang Shoukai Kang Weiwei Xue Zengpeng Li |
| author_facet | Linfei Zhao Mingliang Chen Xiaona Wang Shoukai Kang Weiwei Xue Zengpeng Li |
| author_sort | Linfei Zhao |
| collection | DOAJ |
| description | Tumor necrosis factor α (TNFα), an important clinical testing factor and drug target, can trigger serious autoimmune diseases and inflammation. Thus, the TNFα antibodies have great potential application in diagnostics and therapy fields. The variable binding domain of IgNAR (VNAR), the shark single domain antibody, has some excellent advantages in terms of size, solubility, and thermal and chemical stability, making them an ideal alternative to conventional antibodies. This study aims to obtain VNARs that are specific for mouse TNF (mTNF) from whitespotted bamboosharks. After immunization of whitespotted bamboosharks, the peripheral blood leukocytes (PBLs) were isolated from the sharks, then the VNAR phage display library was constructed. Through phage display panning against mTNFα, positive clones were validated through ELISA assay. The affinity of the VNAR and mTNFα was measured using ELISA and Bio-Layer Interferometry. The binding affinity of 3B11 VNAR reached 16.7 nM. Interestingly, one new type of VNAR targeting mTNF was identified that does not belong to any known VNAR type. To understand the binding mechanism of VNARs to mTNFα, the models of VNARs-mTNFα complexes were predicted by computational modeling combining HawkDock and RosettaDock. Our results showed that four VNARs’ epitopes overlapped in part with that of mTNFR. Furthermore, the ELISA assay shows that the 3B11 potently inhibited mTNFα binding to mTNFR. This study may provide the basis for the TNFα blockers and diagnostics applications. |
| first_indexed | 2024-03-10T03:32:20Z |
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| id | doaj.art-e7081c013d34400eb06feb1bfde88432 |
| institution | Directory Open Access Journal |
| issn | 1660-3397 |
| language | English |
| last_indexed | 2024-03-10T03:32:20Z |
| publishDate | 2022-04-01 |
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| record_format | Article |
| series | Marine Drugs |
| spelling | doaj.art-e7081c013d34400eb06feb1bfde884322023-11-23T11:54:17ZengMDPI AGMarine Drugs1660-33972022-04-0120530710.3390/md20050307Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>)Linfei Zhao0Mingliang Chen1Xiaona Wang2Shoukai Kang3Weiwei Xue4Zengpeng Li5School of Fisheries and Life, Shanghai Ocean University, Shanghai 201306, ChinaKey Laboratory of Marine Genetic Resources, State Key Laboratory Breeding Base of Marine Genetic Resources, Fujian Key Laboratory of Marine Genetic Resources, Fujian Collaborative Innovation Centre for Exploitation and Utilization of Marine Biological Resources, Third Institute of Oceanography Ministry of Natural Resources, Xiamen 361005, ChinaSchool of Pharmaceutical Sciences, Chongqing University, Chongqing 401331, ChinaDepartment of Biochemistry, Institute for Protein Design, University of Washington, Seattle, WA 98195, USASchool of Pharmaceutical Sciences, Chongqing University, Chongqing 401331, ChinaKey Laboratory of Marine Genetic Resources, State Key Laboratory Breeding Base of Marine Genetic Resources, Fujian Key Laboratory of Marine Genetic Resources, Fujian Collaborative Innovation Centre for Exploitation and Utilization of Marine Biological Resources, Third Institute of Oceanography Ministry of Natural Resources, Xiamen 361005, ChinaTumor necrosis factor α (TNFα), an important clinical testing factor and drug target, can trigger serious autoimmune diseases and inflammation. Thus, the TNFα antibodies have great potential application in diagnostics and therapy fields. The variable binding domain of IgNAR (VNAR), the shark single domain antibody, has some excellent advantages in terms of size, solubility, and thermal and chemical stability, making them an ideal alternative to conventional antibodies. This study aims to obtain VNARs that are specific for mouse TNF (mTNF) from whitespotted bamboosharks. After immunization of whitespotted bamboosharks, the peripheral blood leukocytes (PBLs) were isolated from the sharks, then the VNAR phage display library was constructed. Through phage display panning against mTNFα, positive clones were validated through ELISA assay. The affinity of the VNAR and mTNFα was measured using ELISA and Bio-Layer Interferometry. The binding affinity of 3B11 VNAR reached 16.7 nM. Interestingly, one new type of VNAR targeting mTNF was identified that does not belong to any known VNAR type. To understand the binding mechanism of VNARs to mTNFα, the models of VNARs-mTNFα complexes were predicted by computational modeling combining HawkDock and RosettaDock. Our results showed that four VNARs’ epitopes overlapped in part with that of mTNFR. Furthermore, the ELISA assay shows that the 3B11 potently inhibited mTNFα binding to mTNFR. This study may provide the basis for the TNFα blockers and diagnostics applications.https://www.mdpi.com/1660-3397/20/5/307whitespotted bamboosharkIgNARVNARsingle domain antibodyTNFα |
| spellingShingle | Linfei Zhao Mingliang Chen Xiaona Wang Shoukai Kang Weiwei Xue Zengpeng Li Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) Marine Drugs whitespotted bambooshark IgNAR VNAR single domain antibody TNFα |
| title | Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) |
| title_full | Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) |
| title_fullStr | Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) |
| title_full_unstemmed | Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) |
| title_short | Identification of Anti-TNFα VNAR Single Domain Antibodies from Whitespotted Bambooshark (<i>Chiloscyllium plagiosum</i>) |
| title_sort | identification of anti tnfα vnar single domain antibodies from whitespotted bambooshark i chiloscyllium plagiosum i |
| topic | whitespotted bambooshark IgNAR VNAR single domain antibody TNFα |
| url | https://www.mdpi.com/1660-3397/20/5/307 |
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