鸡γ-干扰素与补体受体2蛋白相互作用的研究  

Study on the Interaction Between Chicken Interferonγand Complement Receptor 2 Protein

作　　者：靳换 孟昭英 涂敏 沈佳[1] 史爱华[1] 赵蕾[1] 习硕 张建伟[1] 章振华[1] JIN Huan;MENG Zhaoying;TU Min;SHEN Jia;SHI Aihua;ZHAO Lei;XI Shuo;ZHANG Jianwei;ZHANG Zhenhua(Institute of Animal Husbandry and Veterinary Medicine,Beijing Academy of Agriculture and Forestry Sciences,Beijing 100097,China;Animal Science and Technology College,Beijing University of Agriculture,Beijing 102206,China)

机构地区：[1]北京市农林科学院畜牧兽医研究所,北京100097 [2]北京农学院动物科学技术学院,北京102206

出　　处：《中国畜牧兽医》2024年第10期4476-4484,共9页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金青年基金(32202793);北京市农林科学院创新能力建设(KJCX20230424、KJCX20240338)。

摘　　要：【目的】研究鸡γ-干扰素(chicken interferonγ,chIFN-γ)与鸡补体受体2(chicken complement receptor 2,chCR2)蛋白的相互作用,为研究chIFN-γ和chCR2新功能奠定基础。【方法】通过同源重组方法构建chIFN-γ相关表达质粒,利用免疫共沉淀、激光共聚焦和表面等离子共振技术鉴定chIFN-γ与chCR2蛋白的相互作用,并通过分子对接技术模拟chIFN-γ与chCR2蛋白的相互作用模式。【结果】成功构建了pCMV-Myc-chIFN-γ表达质粒。将pCMV-Myc-chIFN-γ和pCMV-HA-chCR2-ΔTM质粒共转染HEK-293FT细胞,结果表明chIFN-γ蛋白可以与chCR2蛋白进行相互作用;将pCMV-Myc-chIFN-γ和pCMV-HA-chCR2-ΔTM质粒共转染DF-1细胞,结果表明chIFN-γ蛋白与chCR2蛋白可以在DF-1细胞中进行共定位。表面等离子共振试验结果显示chIFN-γ与chCR2蛋白之间的平衡解离常数(KD)值为0.362μmol/L,二者之间有较高的亲和力。分子对接试验结果显示,chIFN-γ与chCR2蛋白之间的相互作用模式为Asp23-His150间形成1组盐桥作用,Asn37-Ser143、Glu57-Gly179和Ile93-Tyr184间形成3组氢键作用。【结论】本研究结果证明了chIFN-γ与chCR2蛋白可以进行相互作用,且二者之间的作用模式为1组盐桥和3组氢键作用。【Objective】The experiment was to study the interaction between chicken interferonγ(chIFN-γ)and chicken complement receptor 2(chCR2)proteins,and lay a foundation for the research of new functions of chIFN-γand chCR2.【Method】chIFN-γrelated expression plasmid was constructed by homologous recombination method.The interaction between chIFN-γand chCR2 proteins was identified by co-immunoprecipitation,laser confocal and surface plasmon resonance techniques,and the interaction mode of chIFN-γand chCR2 proteins was simulated by molecular docking.【Result】The pCMV-Myc-chIFN-γexpression plasmid was constructed successfully.The plasmid of pCMV-Myc-chIFN-γand pCMV-HA-chCR2-ΔTM were co-transfected into HEK-293FT cells and the results showed that chIFN-γcould interact with chCR2.The plasmid of pCMV-Myc-chIFN-γand pCMV-HA-chCR2-ΔTM were co-transfected into DF-1 cells and the results showed that chIFN-γand chCR2 proteins could co-locate in DF-1 cells.The results of surface plasmon resonance test showed that the equilibrium dissociation constant(KD)between chIFN-γand chCR2 proteins was 0.362μmol/L,and the affinity between the two proteins was high.The results of molecular docking experiments showed that the interaction between chIFN-γand chCR2 proteins was 1 salt bridge between Asp23-His150 and 3 sets of hydrogen bonds between Asn37-Ser143,Glu57-Gly179 and Ile93-Tyr184.【Conclusion】The results of this study proved that chIFN-γand chCR2 proteins could interact,and the interaction mode between them was 1 salt bridge and 3 sets of hydrogen bonds.

关 键 词：鸡γ-干扰素(chIFN-γ) 鸡补体受体2(chCR2) 相互作用 亲和力 

分 类 号：S831[农业科学—畜牧学]