Identifying and hijacking tissue-restricted ubiquitin E3 ligase for targeted protein degradation

E3连接酶是催化泛素转移到底物蛋白的多种酶。E3连接酶传统上被认为是对小分子具有挑战性的靶标。由于其复杂的结构特征，以及需要调节蛋白质-蛋白质相互作用，往往涉及较浅的和难以靶向的结合部位(1)。近年来，靶向蛋白降解领域的出现将E3连接酶作为靶标类带到了风口浪尖。E3连接酶可以被称为“降解物”的小分子增选，以招募在正常情况下不是它们的底物的致病蛋白质，导致它们的泛素化和降解。这种小分子包括分子胶降解剂，单价或二价或三价嵌合分子，也被称为靶向嵌合体的蛋白质分解(PROTAC)。小分子结合剂也可以作为E3调节剂，抑制或激活酶，甚至稳定，即提高其蛋白质水平--这允许直接调节E3连接酶本身参与的途径。Ciulli实验室致力于开发E3连接酶靶向配体，包括PROTAC和分子胶。Ciulli实验室率先开发和理解了以结构为导向的E3连接酶配体和针对von Hippel-Lindau(VHL)E3连接酶(2)的PROTAC。Virdee实验室率先开发了用于研究E3连接酶的新型化学生物学工具。这些措施包括开发基于活性的探针，测量泛素结合酶的标志性反硫基化活性，从而深入了解与疾病相关的E3调节(3)。本项目旨在研究新的E3连接酶，并将其与小分子配基。该项目最初将专注于挖掘MS-蛋白质组学数据库，以确定在感兴趣的组织中优先表达的E3连接酶。同时，将开发和应用在Virdee实验室开发的基于活动的探测器，以评估它们的相关活动和功能。一旦满足相对丰度和功能标准的E3连接酶得到验证，我们将使用退化标签，例如我们的BromoTag技术，诱导E3连接酶与组织所需的致病靶蛋白(S)之间的亲和力，以验证小分子诱导的活性。最后，使用基于结构的设计将开发小分子E3粘合剂，这将使PROTAC分子胶的设计成为可能。该项目将解开用于阐明生物学和改进疗法的新型可劫持E3连接酶，学生将在基础和翻译化学和结构生物学的尖端研究方面接受多学科领域的杰出培训。

E3 ligases are diverse enzymes that catalyse the transfer of ubiquitin to substrate proteins. E3 ligases were traditionally viewed to be challenging targets to small molecules. because of their complex structural features, and the need to modulate protein-protein interactions often involving shallow and difficult-to-target binding sites (1). Recently, the advent of the field of "targeted protein degradation" has brought E3 ligases to the forefront as target class. E3 ligases can be co-opted by small molecules called "degraders" to recruit disease-causing proteins that are not their substrates under normal conditions, leading to their ubiquitination and degradation. Such small molecules include "molecular glue" degraders, either monovalent or bi- or tri-valent chimeric molecules, also known as proteolysis targeting chimeras (PROTACs). Small molecules binders can also act as E3 modulators, inhibiting or activating the enzyme, or even stabilizing i.e. increasing its protein level - which allows to modulate directly the pathway in which the E3 ligase is itself involved. The Ciulli lab works on developing E3 ligase targeting ligands including PROTACs and molecular glues. The Ciulli lab has pioneered the structure-guided development and understanding of E3 ligase ligands and PROTACs targeting the von Hippel-Lindau (VHL) E3 ligase (2). The Virdee lab has pioneered the development of novel chemical biology tools for studying E3 ligases. These include the development of activity-based probes that measure the hallmark trans-thiolation activity of ubiquitin conjugation enzymes, allowing insights into disease-relevant E3 regulation (3). This project aims to study new E3 ligases and ligand them with small molecules. The project will initially focus on mining MS-proteomics databases to identify E3 ligases preferentially expressed in tissues of interest. In parallel, activity-based probes developed in the Virdee lab will be developed and applied to assess their relative activities and functionalities. Once E3 ligases meeting criteria of relative abundance and functionalities are validated, we will use degron tags e.g. our BromoTag technology, to induce proximity between the E3 ligase and tissue-required disease-causing target protein(s), to validate small-molecule induced activity. Finally, using structure-based design small-molecule E3 binders will be developed that will enable design of PROTAC molecular glues. The project will untap novel hijackable E3 ligases for illuminating biology and improved therapies and the student will receive outstanding training in multidisciplinary areas in cutting-edge research in fundamental and translational chemical and structural biology.