An integrative structural biology approach to the study of T cell signaling -Equipment Supplement

总结 我建议开发和应用创新的混合结构生物学工具来研究 通过T细胞受体（TCR）复合物进行信号传导的分子机制。这是一 细胞毒性CD8 + T细胞可以通过基本的适应性免疫途径检测到 病毒的存在和体内肿瘤的发展。免疫功能是通过 持续监测抗原递呈细胞中展示的短肽 主要组织相容性复合物（MHC）分子在细胞表面。入门钥匙 是一种多亚基膜蛋白组装体，由克隆型TCR 识别肽-MHC的异源二聚体，以及不变的CD3共受体 一种六聚体，通过血浆在细胞内传递来自细胞表面的激活信号 膜的除了基本的基础科学价值，在原子水平上表征这一过程， 细节具有重要的临床意义，正如大量的 由信号传导组分及其代谢产物的失调引起的免疫缺陷 交互. 尽管进行了大量的功能和结构研究，但阐明了 信号复合体作为一个整体仍然是非常具有挑战性的传统方法，由于 它的大小和动态复杂性。因此，TCR和CD3亚基之间的相互作用 并且信号传导所需的关键构象变化仍然没有完全表征。 考虑到这些瓶颈，我开发了一种新的方法， 互补的方法，如NMR，SANS和cryoEM，以及计算 使用Rosetta程序进行建模，以解决此类具有挑战性的复合物的结构。 在这里，我建议应用这种强大的整合方法来阐明T细胞受体 复杂的结构，并研究其动态转换之间的非活性和活性 构象我的近期目标是确定TCR/CD3相互作用的分子基础 并表征抗原上受体复合物排列的关键结构变化 识别.作为一个长期目标，我计划使用结构引导的方法来设计新颖的T 具有定制特异性和信号传导特性的细胞受体，用于新兴的 免疫疗法的应用。

Summary I propose to develop and apply innovative hybrid structural biology tools to investigate the molecular mechanism of signaling through the T cell receptor (TCR) complex. This is a fundamental adaptive immunity pathway through which cytotoxic CD8+ T cells can detect the presence of viruses and developing tumors in the body. Immune function is achieved through the continuous surveillance of antigen-presenting cells for short peptides displayed within the molecules of the Major Histocompatibility Complex (MHC) on the cell surface. Key to the initiation of signaling, is a multi-subunit membrane protein assembly consisting of a clonotypic TCR heterodimer that recognizes the peptide-MHC, together with the invariant CD3 co-receptor hexamer that relays an activation signal from the cell surface intracellularly, through the plasma membrane. Besides the fundamental basic science merit, characterizing this process at atomic detail has important clinical implications, as is suggested by the large number of immunodeficiencies resulting from dysregulation of the signaling components and their interactions. Despite a large number of functional and structural studies, elucidating the 3D structure of the signaling complex as a whole remains extremely challenging by conventional methods, due to its size and dynamic complexity. As a result, the interactions between the TCR and CD3 subunits and the crucial conformational changes needed for signaling remain incompletely characterized. With these bottlenecks in mind, I have developed a new methodology combining datasets from complementary approaches, such as NMR, SANS and cryoEM, together with computational modeling using the program Rosetta to solve the structures of such challenging complexes. Here, I propose to apply this powerful integrative approach to elucidate the T cell receptor complex structure, and to study its dynamic transitions between inactive and active conformations. My immediate goal is to determine the molecular basis of TCR/CD3 interactions and to characterize the crucial structural changes in receptor complex arrangement upon antigen recognition. As a long-term goal, I plan to use a structure-guided approach to engineer novel T cell receptors with custom specificities and signaling properties, to be used in emerging immunotherapy applications.