| 引用本文: | 束树超,徐明策,哈承旭,周宇杭,邓鹏辉,查丽莎,陈锦霖,陈建明,毕允晨.玻尿酸合成酶的鲨鱼源单域抗体筛选[J].海洋科学,2022,46(8):111-120. |
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| 玻尿酸合成酶的鲨鱼源单域抗体筛选 |
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束树超1,2,3, 徐明策1,2,3, 哈承旭4, 周宇杭5, 邓鹏辉1,2,3, 查丽莎5, 陈锦霖6, 陈建明6, 毕允晨1,2,3
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1.中国科学院海洋研究所, 中国科学院暨山东省实验海洋生物学重点实验室, 中国科学院海洋大科学研究中心, 山东 青岛 266071;2.中国科学院大学, 北京 100049;3.青岛海洋科学与技术试点国家实验室 海洋生物学与生物技术功能实验室, 山东 青岛 266237;4.青岛海洋科技馆, 山东 青岛 266003;5.安徽农业大学, 安徽 合肥 230036;6.闽江学院海洋研究院, 福建 福州 350108
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| 摘要: |
| 在鲨鱼等软骨鱼类体内存在天然的缺失轻链、仅包含重链的抗体,源于这种重链抗体可变区的片段称为单域抗体(single domain antibody)。目前单域抗体研究中的抗原主要来源于水溶性蛋白或病原体,筛选膜蛋白鲨鱼源单域抗体的领域接近空白。本研究以重组表达的小球藻病毒玻尿酸合成酶(一种膜蛋白)为抗原,经过免疫、建库、淘选、验证等步骤获得了抗原特异性条纹斑竹鲨(Chiloscyllium plagiosum)单域抗体序列。随后利用大肠杆菌表达了该单域抗体,通过等温滴定量热技术(ITC)测定了其与抗原的亲和力,证实了以膜蛋白作为抗原制备鲨鱼源单域抗体的可行性。 |
| 关键词: 单域抗体 纳米抗体 玻尿酸合成酶 |
| DOI：10.11759/hykx20210118001 |
| 分类号:Q816 |
| 基金项目:国家自然科学基金(41876168);青岛海洋科学与技术试点国家实验室海洋生物学与生物技术功能实验室引进人才支持计划(YJ2019NO03);中科院合肥大科学中心高端用户培育基金(2020HSC-UE018) |
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| Screening of shark-derived single domain antibodies for hyaluronan synthase |
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HU Shu-chao1,2,3, XU Ming-ce1,2,3, HA Cheng-xu4, ZHOU Yu-hang5, DENG Peng-hui1,2,3, ZHA Li-sha5, CHEN Jin-lin6, CHEN Jian-ming6, BI Yun-chen1,2,3
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1.CAS and Shandong Province Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Center for Ocean Mega-Science, Chinese Academy of Sciences, Qingdao 266071, China;2.University of Chinese Academy of Sciences, Beijing 100049, China;3.Marine Biology and Biotechnology Laboratory, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao 266237, China;4.Ocean Science and Technology Museum of Qingdao, Qingdao 266003, China;5.Anhui Agricultural University, Hefei 230036, China;6.Institute of Oceanography Minjiang University, Fuzhou 350108, China
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| Abstract: |
| The single domain antibody (sdAb) is derived from the variable fragment of the heavy-chain-only antibody, which has been found naturally in cartilaginous fishes, such as sharks. The sdAb research mainly arise from the soluble proteins and pathogens as the antigens to date. In this study, after the steps of immunogen preparation, immunization, repertoire cloning, library construction, selection and verificatione obtained the shark-derived sdAb, which specifically recognizes Chlorella virus hyaluronan synthase (a membrane protein). The sdAb was then expressed in E.coli and purified for the binding affinity determination performed by Isothermal Titration Calorimetry technology (ITC). Overall, this study proves the practicability of generating shark-derived single-domain antibodies against membrane protein antigen. |
| Key words: single domain antibody nanobody hyaluronan synthase |
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