Structure Of Complement C3(H2O) Revealed By Quantitative Cross-Linking/Mass Spectrometry And Modeling

The slow but spontaneous and ubiquitous formation of C3(H2O), the hydrolytic and conformationally rearranged product of C3, initiates antibody-independent activation of the complement system that is a key first line of antimicrobial defense. The structure of C3(H2O) has not been determined. Here we subjected C3(H2O) to quantitative cross-linking/mass spectrometry (QCLMS). This revealed details of the structural differences and similarities between C3(H2O) and C3, as well as between C3(H2O) and its pivotal proteolytic cleavage product, C3b, which shares functionally similarity with C3(H2O). Considered in combination with the crystal structures of C3 and C3b, the QCMLS data suggest that C3(H2O) generation is accompanied by the migration of the thioester-containing domain of C3 from one end of the molecule to the other. This creates a stable C3b-like platform able to bind the zymogen, factor B, or the regulator, factor H. Integration of available crystallographic and QCLMS data allowed the determination of a 3D model of the C3(H2O) domain architecture. The unique arrangement of domains thus observed in C3(H2O), which retains the anaphylatoxin domain (that is excised when C3 is enzymatically activated to C3b), can be used to rationalize observed differences between C3(H2O) and C3b in terms of complement activation and regulation. <strong>For more information</strong> about how to reproduce this modeling, see the Sali lab website or the README file.缓慢、自发且广泛存在的C3(H2O)——C3经水解与构象重排得到的产物——的形成，会启动补体系统的抗体非依赖性激活，而补体系统是抗菌防御的关键第一道防线。目前尚未解析C3(H2O)的晶体结构。本研究对C3(H2O)开展了定量交联质谱法（quantitative cross-linking/mass spectrometry, QCLMS）分析。该分析揭示了C3(H2O)与C3，以及C3(H2O)与其关键蛋白水解裂解产物C3b之间的结构异同，其中C3b与C3(H2O)存在功能相似性。结合C3与C3b的晶体结构，QCLMS数据表明，C3(H2O)的生成伴随C3的含硫酯结构域从分子的一端迁移至另一端。这一过程会形成一个稳定的C3b样平台，可结合酶原B因子（factor B）或调节因子H因子（factor H）。整合现有晶体学与QCLMS数据，我们得以构建C3(H2O)的结构域架构三维模型。我们在C3(H2O)中观察到的独特结构域排布——其保留了过敏毒素结构域（该结构域在C3经酶促激活为C3b时会被切除）——可以合理解释C3(H2O)与C3b在补体激活与调控方面的差异。<strong>如需获取更多相关信息</strong>：若需复现该建模流程，请参阅Sali实验室官网或README文件。