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| 基于表面等离子共振技术鉴定鸡补体受体2与其配体chC3d的亲和力 |
| 靳换,杨芊芊,王聪,涂敏,赵蕾,澎湃,王海良,孟昭英,李永清,章振华 |
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| (北京市农林科学院畜牧兽医研究所) |
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| 摘要: |
| 鸡补体受体2(Chicken complement receptor 2, chCR2)是一种表达于鸡B淋巴细胞细胞表面的跨膜蛋白,chCR2可以与其生理学配体鸡补体组分3d(Chicken complement component 3d, chC3d)相互作用。目前评估受体与配体亲和力的方法主要有表面等离子共振技术(Surface plasmon resonance, SPR)和生物膜干涉技术(Biolayer interferometry, BLI)等。本研究旨在利用表面等离子共振技术鉴定chCR2受体分子与其配体chC3d分子的亲和力。本研究成功构建了重组真核表达载体pTT5-chCR2和pTT5-chC3d；利用HEK 293F表达系统表达了His-chCR2和His-chC3d蛋白；利用NI柱纯化系统纯化了His-chCR2和His-chC3d蛋白；利用表面等离子共振技术,通过分析chCR2与chC3d结合的动力学参数得出两者之间的平衡解离常数KD为1.37μmol/L。结果表明chCR2与其配体chC3d以高亲和力结合。 |
| 关键词: 鸡补体受体2：鸡补体组分3d 表面等离子共振技术 相互作用。 |
| DOI： |
| 投稿时间：2023-04-23修订日期：2023-06-16 |
| 基金项目:国家自然科学基金（32202793）；北京市农林科学院青年基金（QNJJ202111）；北京市农林科学院畜牧兽医研究所改革与发展（XMS202313）；北京市农林科学院创新能力建设（KJCX20230424）。 |
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| The identification of binding affinity between chicken compliment receptor 2 and its ligands chC3d by surface plasmon resonance |
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| (Beijing Academy of Agriculture and Forestry Science) |
| Abstract: |
| Chicken complement receptor 2 (chCR2) is a transmembrane protein expressed on the surface of chicken B lymphocytes. chCR2 can interact with its physiological ligand chicken complement component 3d (chC3d). At present, the methods to evaluate the affinity between receptor and ligand mainly include Surface plasmon resonance (SPR) and biomembrane interference (BLI). In this study，the SPR was to identify the affinity between the chCR2 and its ligand chC3d. The recombinant eukaryotic expression vectors pTT5-chCR2 and pTT5-chC3d were successfully constructed ; His-chCR2 and His-chC3d proteins were expressed using the HEK 293F expression system; The His-chCR2 and His-chC3d proteins were purified using the NI column purification system; The equilibrium dissociation constant KD between chCR2 and chC3d is 1.37 μ mol/L by analyzing the kinetic parameters of the combination of chCR2 and chC3d. The results indicate that chCR2 binds to its ligand chC3d with high affinity. |
| Key words: Chicken complement receptor 2,Chicken complement component 3d,Surface plasmon resonance,Interaction |
