Novel assay for ubiquitin pathway enzymes as biomarkers

描述(申请人提供)：众所周知，泛素途径在神经生物学中起着重要的调节作用。泛素途径酶是成熟神经元突触适应性的重要调节因子，功能障碍的泛素途径酶或调节与多种神经退行性疾病有关。脱泛素酶，或DUBS属于一个约100个主要是单一亚基的蛋白质家族，它能从目标蛋白质的赖氨酸残基中去除泛素。有证据表明，调控失调或不调控的Dub(S)会导致神经退行性疾病。大多数常用的DUB活性检测方法既不适合临床应用，也不适合药物开发。在这项提案中，将开发一种新的DUB蛋白家族的分析方法。该分析涉及一种半合成方法来开发试剂，该试剂是连接到目标蛋白的泛素的C末端的延伸。多肽文库可以从泛素的C末端延伸而来；对于给定的DUB，可以从文库中选择最佳试剂，因为DUB的类别显示出不同的特异性偏好。与它们在溶液相分析中的命运相反，被DUBS切割的实体将被固定在表面上，使它们能够在多重环境中作为固定DUB的合适试剂。由于该检测可以识别出一种DUB蛋白，它还可以发现DUB相互作用蛋白或神经退行性疾病中常见的泛素化底物积累。因此，该分析将用于诊断和药物发现目的。在第二阶段，化验将扩大到包括所有已知的DUB。这项技术的商业应用是用于诊断和治疗的产品的产生，用于与配音相关的医疗需求，如神经退行性疾病。被另一种称为泛素的小蛋白修饰的蛋白质是神经生物学的关键调节因子。催化泛素在靶蛋白上的添加或移除的酶是神经细胞功能的重要调节因子，在诊断各种神经系统疾病或发现治疗这些疾病的药物方面具有潜在的应用价值。与公共健康相关：Lifesensors建议开发一种检测泛素途径酶的主要类型，称为DUBS(脱泛素酶)。这项测试代表了一项新技术，它将在诊断和发现治疗阿尔茨海默氏症和其他神经疾病的有效和选择性药物方面具有潜在的实用价值。

DESCRIPTION (provided by applicant): It has been well established that the ubiquitin pathway plays an important role as a key regulator of neuronal biology. The ubiquitin pathway enzymes are essential and critical mediators of synaptic adaptability in mature neurons, and dysfunctional ubiquitin pathway enzymes or regulation is associated with diverse neurodegenerative disorders. Deubiquitinating enzymes, or DUBs belong to a family of ~100 primarily single subunit proteins that remove ubiquitin from lysine residues of target proteins. Evidence suggests that that dysregulated or un-regulated DUB(s) can lead to neurodegenerative disease. Most commonly used assays for DUB activity are not suitable for either clinical applications or drug discovery. In this proposal, a novel assay for the DUB family of proteins will be developed. The assay involves a semi-synthetic approach to develop reagents that are extensions of the C-terminus of ubiquitin that is conjugated to target proteins. A library of peptides can be extended from the C-terminus of ubiquitin; the best reagent for a given DUB can be selected from the library since classes of DUBs show different specificity preferences. In contrast to their fate in solution phase assays, entities cleaved by DUBs will be immobilized on a surface, allowing them to be suitable reagents for immobilized DUBs in a multiplex environment. Since the assay can identify a protein as a DUB, it can also uncover DUB interactive proteins or the accumulation of ubquitinated substrates common in neurodegenerative diseases. Thus, the assay will be useful for diagnostic as well as drug discovery purposes. In Phase II, the assay will be expanded to include all known DUBs. The commercial application of this technology is the generation of products for diagnostic and therapeutic uses as applied to DUB-linked medical needs such as neurodegenerative diseases. Proteins modified by another small protein called ubiquitin are key regulators of neuronal biology. Enzymes that catalyse the addition or removal of ubiquitin to target proteins are important regulators of neuronal cell function and are potentially useful for diagnosing various neurological diseases or discovering drugs to treat the diseases. PUBLIC HEALTH RELEVANCE: LifeSensors proposes to develop an assay for one of the major types of ubiquitin pathway enzymes known as DUBs (deubiquitinating enzymes). This assay represents a novel technology that will have potential utility in diagnostics and in the discovery of potent and selective drugs to treat Alzheimer's Disease and other neurological diseases.