Dynamics of transmembrane dimers in TNF-Receptors by EPR and molecular simulation

描述（由申请人提供）：死亡受体5 （DR5）是跨膜受体tnf超家族的成员，在凋亡信号通路中起关键作用。DR5在癌细胞中表达上调，是临床和基础研究中最积极追求的抗癌靶点之一。然而，为了最大限度地促进癌细胞的凋亡，迫切需要开发新的靶向DR5的策略。要做到这一点，我们需要更多的结构和生物物理数据，以了解受体是如何工作的。传统上，研究主要集中在细胞外配体结合域的晶体结构上。因此，在信号转导过程中，关于蛋白质跨膜区域内发生的关键结构事件的数据仍然非常缺乏。最近，多项高影响力的研究表明，了解配体诱导的跨膜结构域的结构和动力学变化？-螺旋二聚体是了解受体功能的下一个关键步骤。该应用程序的目的是了解与受体活性和非活性状态相关的DR5跨膜结构的关键变化，并确定关键氨基酸基序

DESCRIPTION (provided by applicant): Death Receptor 5 (DR5) is a member of the TNF-superfamily of transmembrane receptors that plays a critical role in signaling the apoptotic pathway. Upregulated in cancer cells, DR5 is among the most actively pursued anti-cancer targets, both clinically and in basic research studies. However, there is great need to develop new strategies for targeting DR5 in order to maximize apoptosis in cancer cells. To do so, we need significantly more structural and biophysical data in order to understand how the receptor works. Traditionally, research has focused on the crystal structure of the extracellular, ligand-binding domain. As such, there remains a debilitating scarcity of data regarding the key structural events that occur within the transmembrane domain of the protein during transduction of the signal. Very recently, multiple high impact studies have shown that understanding the ligand-induced changes in the structure and dynamics of the transmembrane domain ?- helical dimer is the next crucial step in understanding the function of the receptor. The objective of thi application is to understand key changes in the transmembrane structure of DR5 associated with the active and inactive states of the receptor and to determine the critical amino-acid motifs that dictate changes in conformation. The rationale of this proposal is that once we understand important conformational states of the DR5 TM domain, and the most relevant motifs that stabilize states of the protein, we will be able to evaluate its potential as a therapeutic target or pharmacological regulation. Our approach combines molecular biophysics experiments on model systems (synthetic TM domains) complemented by computational simulations and molecular biology experiments on full-length receptors in living cells. We will 1) define the inter helical architecture of the DR5 transmembrane domain dimer; 2) identify key sequence alterations that either disrupt TM dimerization or stabilize alternate dimer conformations; and 3) establish the capacity to modulate TM dimer architecture and affect DR5 activation in cancer cells. The proposed research will advance understanding of TNF-Receptors in general, taking the logical but critical next steps in building a complete description of their structure-function relationship. We will create new understanding of the physical principles that dictate conformational dynamics of TM dimers, principles that are essential in a broad range of membrane protein superfamilies. In the process, we will advance the state-of-the- art in computational modeling of membrane proteins, providing a methodological roadmap for validation of models by comparison to experimental EPR spectroscopy. Working with a pancreatic cancer researcher will enable us to evaluate the TM domain of DR5 as a target for future therapeutic intervention.