Targeted degradation of proteins by affinity peptide conjugated ubiquitin (APCU)

亲和肽偶联泛素(APCU)对蛋白质的靶向性降解 蛋白质降解具有许多DNA/RNA操作无法覆盖的独特特性 工具。为了了解生物过程的机制，并达到理想的治疗效果， 目前迫切需要旨在诱导蛋白质降解的技术。然而，很少有选项是 目前可用于此类目的。此外，现有的工具也有已知的局限性，例如难以 适应或钟形剂量-反应曲线，或易受De-De负调节 泛素化系统。为了扩大工具的选择范围，并克服当前方法的局限性，我们 建议建立一种新的方法来诱导蛋白质降解，方法是将泛素部分与 与目标蛋白质有高亲和力的分子。对于证明或概念，将首先使用多肽作为亲和力 分子。因此，这个原型被称为亲和肽结合泛素(APCU)。在我们的初步研究中， 我们设计了一个带有针对MCL1的多肽的APCU分子原型，MCL1是一种过度表达的癌蛋白 在许多类型的癌症中。我们发现，这种分子可以通过以下方式特异性地降低MCL1蛋白的水平 促进它的退化。在拟议的研究中，我们将确定APCU的行动机制 方法和设计这类分子的最佳条件。我们还将确定剂量 MCL1靶向APCU的响应曲线及其对去泛素化系统的敏感性。此外，要 为APCU的广泛应用和未来的活体应用做准备，我们将在其他几个平台上测试APCU的应用 蛋白质，并探索用条件结合策略形成APCU分子的可行性。这 拟议中的研究，如果脱离，将提供一个非常有价值的工具，专门降解蛋白质的机械 调查和潜在的治疗用途。

Targeted degradation of proteins by affinity peptide conjugated ubiquitin (APCU) Protein degradation possesses many unique properties that cannot be covered by DNA/RNA manipulation tools. To understand the mechanism of biological processes, and to achieve desirable therapeutic effects, techniques designed to induce protein degradation are in critical needs. However, very few options are currently available for such purposes. Also, the existing tools have known limitations such as difficulty for adaptation or bell-shaped dose-response curve, or vulnerability for a negative regulation by the de- ubiquitination system. To expand the option for tools, and to overcome limitations of current methods, we propose to establish a new approach to induce protein degradation by conjugating a ubiquitin moiety with a molecule of high affinity to a target protein. For a proof or concept, peptides will first be used as the affinity molecule. This prototype is thus called affinity peptide conjugated ubiquitin (APCU). In our preliminary study, we designed a prototype APCU molecule with a peptide targeting MCL1, an oncoprotein that is over-expressed in many types of cancer. We found that this molecule can specifically reduce the level of MCL1 protein by promoting its degradation. In the proposed study, we will determine the action mechanism of the APCU approach and the optimal conditions for designing this types of molecules. We will also determine the dose response curve of the MCL1-targeting APCU and its sensitivity of de-ubiquitination system. Furthermore, to prepare for a wide usage and future in vivo applications, we will test the application of APCU on several other proteins and explore the feasibility of forming an APCU molecule with a conditional conjugation strategy. This proposed study, if secedes, will provide a highly valuable tool to specifically degrade a protein for mechanistic investigations and for potential therapeutic usages.