Zhong Chen, Huo Rongfen, Hu Kuan, Shen Jinlong, Li Dangsheng, Li Ningli, Ding Jianping
National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, Center for Excellence in Molecular Cell Science, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
Shanghai Institute of Immunology and Department of Immunology and Microbiology, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
J Mol Recognit. 2017 Nov;30(11). doi: 10.1002/jmr.2645. Epub 2017 Jun 13.
CCN1, also named Cyr61 (cysteine-rich protein 61), is the first identified member of the CCN family that is composed of 6 secreted extracellular matrix-associated glycoproteins. CCN1 has been demonstrated to participate in pathogenesis of rheumatoid arthritis through various pathways. A monoclonal antibody, namely, 093G9, is effective to antagonize the effects of CCN1 and hence has potential therapeutic benefits against rheumatoid arthritis. Here, we show that the epitope recognized by 093G9 is mapped to residues 77 to 80 of CCN1, and a cyclic peptide encompassing residues 75 to 81 of CCN1 displays high binding affinity for 093G9. The crystal structure of the 093G9 Fab in complex with the cyclic peptide was determined at 2.7 Å resolution, which reveals the intensive interactions between CCN1 and 093G9. Particularly, residues Asn79 and Phe80 of CCN1 are inserted into cavities mainly formed by residues of complementarity-determining region loop L3 and framework region L2 and by residues of complementarity-determining region loops H2 and H3, respectively, which contribute most of the interactions and therefore are critical for the recognition by 093G9. Together, these findings not only identify the epitope of CCN1 for 093G9 but also reveal the molecular mechanism of recognition and binding of CCN1 by 093G9.
CCN1，也被称为Cyr61（富含半胱氨酸的蛋白质61），是CCN家族中第一个被鉴定出的成员，该家族由6种分泌型细胞外基质相关糖蛋白组成。CCN1已被证明通过多种途径参与类风湿性关节炎的发病机制。一种单克隆抗体，即093G9，可有效拮抗CCN1的作用，因此对类风湿性关节炎具有潜在的治疗益处。在此，我们表明093G9识别的表位定位于CCN1的第77至80位残基，并且一个包含CCN1第75至81位残基的环肽对093G9具有高结合亲和力。093G9 Fab与环肽复合物的晶体结构在2.7 Å分辨率下确定，这揭示了CCN1与093G9之间的紧密相互作用。特别地，CCN1的Asn79和Phe80残基分别插入到主要由互补决定区环L3和框架区L2的残基以及互补决定区环H2和H3的残基形成的腔中，这些残基贡献了大部分相互作用，因此对于093G9的识别至关重要。总之，这些发现不仅确定了CCN1对093G9的表位，还揭示了093G9对CCN1识别和结合的分子机制。
