| 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（平衡解离常数约为171 μM），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： |
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| 基金项目:国家自然科学基金项目（面上项目，重点项目，重大项目），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 ， 中科院合肥大科学中心高端用户培育基金 |
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| Screening of shark-derived Single Domain Antibodies for Hyaluronan Synthase |
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SHU Shuchao1, XU Mingce1, ZHOU Yuhang2, DENG Penghui1, ZHA Lisha2, CHEN Jinlin3, CHEN Jianming3, BI Yunchen1
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1.Institute of Oceanology, Chinese Academy of Sciences;2.Anhui Agricultural University;3.Institute of Oceanography Minjiang University
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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, we obtained the shark-derived sdAb, which specifically recognizes Chlo-rella 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 Calorime-try technology (ITC). The equilibrium dissociation constant is approximately 171 μM. 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 |