中东呼吸综合征冠状病毒（Middle East respiratory syndrome coronavirus，MERS-CoV）于2012年在中东地区首次被鉴定出来，随后逐渐扩散，在很多国家都报道了感染病例。MERS-CoV能导致非常严重的呼吸道病症，致死率很高，引发了全球关于MERS大流行潜在可能性的广泛关注。人的细胞表面蛋白二肽基肽酶4（Dipeptidyl Peptidase-4，DPP4）是MERS-CoV的受体。MERS-CoV通过其表面S蛋白与DPP4结合，进而攻击人体细胞。本论文首先鉴定了MERS-CoV受体结合区（receptor binding domain，RBD）在S蛋白上的精确位置，并用蛋白质晶体学方法解析了RBD与DPP4复合物的3.0 ?的结构。结果显示，与SARS-CoV RBD类似，MERS-CoV的RBD可以进一步分为两个部分，即骨架区域（core subdomain）和受体结合区域（receptor-binding subdomain）。其中骨架区域在SARS-CoV和MERS-CoV中非常保守，而受体结合区域则差异明显。RBD结合受体DPP4的区域位于DPP4的β-螺旋桨结构域（β-propeller domain），而非α/β水解酶结构域（α/β hydrolase domain）。然后，本论文采用点突变和常用的生化方法（如Pull-down，SPR等），鉴定了RBD和DPP4上对二者的结合起关键作用的氨基酸。同时与清华大学医学院张林琦教授实验室合作，构建了MERS-CoV假病毒系统，进一步验证了这些关键氨基酸位点的突变对假病毒侵染能力的影响。DPP4上的这些关键氨基酸位点在不同物种中的保守性将有助于解释MERS-CoV的宿主选择性。最后，张林琦教授实验室利用我们鉴定的RBD片段，从酵母展示的人源scFv文库中筛选到了两株高中和能力的单克隆抗体。本论文基于此制备了两个中和抗体的Fab片段，然后分别获得了Fab-RBD复合物的晶体。未来的工作将集中于解析这两种晶体的结构，更深入的了解抗体识别的表位及抗体发挥中和作用的机制。RBD-DPP4复合物的晶体结构提供的原子分辨率的信息详细解释了病毒识别受体的机制，这些信息将有助于研发针对MERS-CoV的特异性的预防和治疗方法。而RBD及RBD的抗体有望在将来应用于针对MERS-CoV的特异的预防和治疗，在临床上有很好的应用前景。

The newly emerged Middle East respiratory syndrome coronavirus (MERS-CoV) was firstly identified in humans in the Middle East in September, 2012, and later in several other countries. MERS patients usually develop severe pulmonary illness and the fatality rate is high, which leads to global concern about the potential for a MERS pandemic. Dipeptidyl peptidase 4 (DPP4) is the cellular receptor of MERS-CoV. The binding between the MERS-CoV S protein and human DPP4 mediates viral attachment to host cells and then initiates infection through membrane fusion. We firstly identified the receptor-binding domain (RBD) on the viral S protein, which directly mediates the interaction with DPP4. We then solved the 3.0 ? resolution crystal structure of MERS-CoV RBD complexed with the extracellular domain of human DPP4. Our results show that viral RBD is composed of a core subdomain and a unique receptor-binding subdomain. The receptor-binding subdomain interacts with DPP4 β-propeller domain, instead of its intrinsic α/β hydrolase domain. MERS-CoV RBD and related SARS-CoV RBD have a high structural conservation in their core subdomains, but a notably divergent in the receptor-binding subdomains. Using site specific mutation and other biochemical methods such as pull-down assay and Surface Plasmon Resonance（SPR）, several key residues in the MERS-CoV RBD and in DPP4 protein were identified that are critical for viral binding to DPP4. Besides, we collaborated with Professor Zhang Linqi, developed and utilized a pseudotyped virus system for studying MERS-CoV S protein-mediated infection. With the help of the pseudotyped virus system, key residues at the interface between MERS-CoV RBD and DPP4 were additionally confirmed. Conservation of DPP4 residues that contact the RBD of MERS-CoV among different mammals may hence help to track the animal source for MERS-CoV.Two RBD-specific potent human neutralizing monoclonal antibodies (MERS-4 and MERS-27) were then isolated from yeast-displayed single-chain variable region fragments (scFvs) of nonimmune human antibody library in Zhang Linqi’s lab. To learn the detailed neutralization mechanism of the two mAbs needs the fine-mapping of epitopes and Structural Biology may tell us a lot. We expressed, purified the two antibodies and cutted both of them to generate Fabs. For both MERS4 and MERS27, we got the Fab-RBD complexes and subsequently obtained crystals. Future work would be required to solve the crystal structures. The detailed structural information at the interface between MERS-CoV RBD and DPP4 provides a more complete understanding of the viral attachment to host cells, which can guide development of therapeutics and vaccines against MERS-CoV infection. Besides, RBD and RBD-specific neutralizing antibodies are both promising candidates for prophylactic and therapeutic interventions against MERS-CoV infection.