MIF与CD74复合物结构及其在CD74依赖的信号转导中的作用与机制

MIF plays an important role in the inflammation and cell proliferation, and is involved in the pathogenesis of a lot of diseases and cancers thus becomes an important biomarker and drug target. MIF could bind to its cognate receptor CD74 to initiate the downstream signaling cascade reaction to exert different kinds of biological functions. However, the structural basis of the interaction between MIF and CD74 and the mechanism is still not clear. We have purified recombinant MIF and CD74 and have confirmed that they interact. We will determine the structures of MIF complexed to sCD74 and the ternary complex of specifc compound-MIF-sCD74, identify the key sites for their interactions and conformational changes and the stabilization of the conformations, reveal the roles of MIF catalytic sites and the extracellular region and intracellular domain in signaling through CD74, and elucidate the structural basis for the regulation of specific compounds on the MIF-CD74 complex. This project will clarify the structural basis of the interaction between MIF and CD74, the role and molecular mechanism of MIF in CD74-dependent signal transduction, and lay the foundation for revealing the mechanism of MIF in the the pathogenesis of the diseases and cacers, and provide critical parameters and theorectical instructions for the design and optimization of the drugs targeted for MIF.

MIF在炎症和细胞增殖等信号转导中发挥着重要作用，参与多种疾病与癌症的发生，是重要的生物标记分子和药物标靶。MIF可结合CD74，启动下游信号级联反应，发挥生物学效应。但MIF与CD74相互作用结构基础和机制尚不明确。我们前期已经重组表达纯化MIF和CD74并已证实它们相互作用。本项目将测定MIF-sCD74复合物以及特异性化合物-MIF-sCD74复合物的三维结构，揭示MIF和CD74相互作用和构象变化及稳定的关键位点，鉴定MIF催化位点、CD74的胞外和胞内区域在CD74信号转导中的作用，并明了特异性化合物调控MIF-CD74 复合物的结构基础。本课题将阐明MIF与CD74相互作用的结构基础、MIF在CD74依赖的细胞信号转导中的作用与分子机制，为揭示MIF在疾病和癌症发生中的作用机制奠定基础，并为MIF作为靶标的药物设计提供关键参数和理论。

本研究表达和纯化了MIF和sCD74，开展后续的结晶学实验以及细胞生物学实验。通过已知的MIF的结构进行虚拟筛选化合物，并根据MIF的酶学实验进行初步筛选，并测定多个MIF和鉴定的化合物复合物的晶体结构，并筛选出对MIF与CD74相互作用有显著效果的化合物，检测了化合物对MIF介导巨噬细胞迁移的影响，以及通过CCK-8试剂检测化合物对RAW264.7巨噬细胞活性的影响，综合分析揭示MIF与化合物相互作用的细节与分子机制，明确MIF酶催化位点和CD74胞外区域在生物活性中的作用，为MIF作为靶标的药物设计提供理论基础。随后结合其他文献分析，设计了两种CD74多肽，检测其对MIF介导巨噬细胞迁移的抑制效果以及通过CCK-8试剂检测两种多肽对RAW264.7巨噬细胞活性的影响。然而经过多种尝试均没有获得MIF-sCD74或者MIF-sCD74-化合物复合物的晶体结构。在后续的研究中将要尝试更多的技术，力图获得MIF-sCD74复合物晶体和结构以及MIF-sCD74-化合物晶体和结构。