Project 3: Structural Basis for grp94 Drug Development and Chaperone Function

项目总结 Grp94是一种ER HSP90伴侣，其广泛的致癌作用直到最近才被确定。 然而，对grp94结构和机制的了解一直落后于其他HSP90。GRP94 与其他细胞HSP90类似物显示出高度的序列和结构同源性。尽管如此，尝试 为了证明胞浆和内质网之间的互补在两者中都是持续失败的 方向，表明维持高水平的专门内质网的巨大代谢负担- 特定的HSP90具有明显的机械目的。我们认为，这种独特的机制反映在其 结构。本项目的目标是了解grp94在hsp90家族中的独特之处，并利用 这些特点在设计新的抑制剂作为癌症治疗药物。我们将在四个月内解决这个问题 明确的目标。第一个是表征和开发下一代grp94选择性抑制剂。与.一起 加布里埃拉·奇奥西斯博士，项目2的负责人，我们已经表征了一组基于嘌呤的(PU)化合物 选择性地与grp94结合并显示抗癌活性。亲和力更高，选择性更强的化合物 需要提高它们的治疗潜力，并作为grp94功能的化学探针。第二个是 确定PU配体与grp94选择性结合的结构基础。我们将调查 促进选择性抑制物结合的grp94的构象变化往往发生在一个平行对数中，而不是 另一个--是什么特定的序列和结构元素使grp94反应与众不同？第三，我们的目标是 确定单个结构域如何对细胞的独特生化和功能特性做出贡献 Gp94。我们将与项目1的负责人李子海博士合作，研究单个grp94域是如何 有助于该Paralog的整体专业化和功能，因为它成熟其客户蛋白质，如 加普。第四，我们将探讨翻译后修饰(PTM)在grp94客户端结合和 活动。总之，这些实验应该为开发新的抗癌疗法铺平道路。 并阐明了有关grp94作用的中心力学问题。

PROJECT SUMMARY Grp94 is an ER HSP90 chaperone whose broad oncogenic roles have only recently been established. However, understanding of grp94 structure and mechanism has lagged behind that of other HSP90s. grp94 exhibits high sequence and structural homology to the other cellular HSP90 paralogs. Nevertheless, attempts to demonstrate complementation between the cytosolic and ER paralogs have consistently failed in both directions, suggesting that the enormous metabolic burden of maintaining high levels of a specialized ER- specific HSP90 has a distinct mechanistic purpose. We propose that this distinct mechanism is reflected in its structure. The goal of this project is to understand what makes grp94 unique in the HSP90 family and to exploit these features in the design of new inhibitors as cancer therapeutics. We will address this question in four specific aims. The first is to characterize and develop next-generation grp94-selective inhibitors. Together with Dr. Gabriela Chiosis, leader of Project 2, we have characterized a set of purine-based (PU) compounds that bind selectively to grp94 and exhibit anti-cancer activity. Higher affinity and more selective compounds are needed to improve their therapeutic potential and to serve as chemical probes of grp94 function. The second is to determine the structural basis for selective binding of PU ligands to grp94. We will investigate how the conformational changes in grp94 that foster selective inhibitor binding manage to occur in one paralog and not another – what specific sequence and structural elements make the grp94 response distinct? Third, we aim to determine how individual structural domains contribute to the unique biochemical and functional properties of grp94. In collaboration with Dr. Zihai Li, leader of Project 1, we will examine how individual grp94 domains contribute to the overall specialization and function of this paralog as it matures its client proteins such as GARP. Fourth, we will probe the role of post-translational modifications (PTMs) on grp94 client binding and activity. Together these experiments should pave the way for the development of new anti-cancer therapeutics and illuminate central mechanistic questions about grp94 action.