Functions of Ub-like Proteins and Proteases

Ub 样蛋白和蛋白酶的功能

• 批准号: 7150820
• 负责人: KEITH D WILKINSON
• 金额: $ 32.51万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7150820
• 关键词: catalyst; chemical conjugate; chemical kinetics; chemical synthesis; endopeptidases; enzyme activity; enzyme mechanism; enzyme substrate; gene expression; high throughput technology; protease inhibitor; protein structure function; proteomics; ubiquitin

DESCRIPTION (provided by applicant): We have developed specific inhibitors of, and substrates for, deubiquitinating enzymes (DUBs) acting upon ubiquitin, NeddS, SUMO-1, SUMO-2/3, and ISG15 (collectively referred to as ubiquitin-like or UBL proteins). We refer to these as the UBL-X panel of reagents. The specificity of these reagents is outstanding and methods to assess cross reactivity between DUB homologs and UBL paralogs have been developed. These reagents have allowed us to define the likely specificity of a number of individual DUBs, led to some surprising conclusions regarding the spectrum of activities exhibited by different classes of DUBs, and suggested new opportunities for developing and characterizing inhibitors that specifically interfere with selected pathways of UBL protein metabolism. In the coming period: We will use the UBL-X panel of reagents in quantitative labeling and kinetic approaches to determine the amounts and paralog specificity of active desumoylating enzymes in lysates from control and stressed cells. The specificity will be compared to that of the purified catalytic domains. We will test the hypothesis that changes in cellular levels of sumoylated proteins in response to cellular stress are controlled by levels of active desumoylating enzymes. We will determine the enzymatic specificity of the DUB activity exhibited by the SARS virus papain-like protease using the panel of UBL-X reagents. This information will suggest possible cellular substrates and be used to mount a high throughput drug screen to identify antiviral agents that may be useful in limiting the spread of coronavirus infections. We will use the panel of UBL-X reagents to evaluate the specificity of inhibitors that have been reported to inhibit deubiquitinating enzymes. This will provide a rapid and selective method to evaluate the global effects of "specific" inhibitors on the spectrum of deubiquitinating enzymes. In the broadest sense, these studies will contribute to our understanding of UBL protein metabolism and guide development of inhibitors and drugs for the treatment of cancers and microbial infections.

描述（由申请人提供）：我们已经开发了作用于泛素，NEDDS，SUMO-1，SUMO-2/3和ISG15的去泛素化酶（DUBS）的特定抑制剂（DUBS）（集体称为泛素蛋白样或UBL蛋白）。我们将其称为UBL-X试剂面板。这些试剂的特异性是出色的，并且已经开发了评估配音同源物和UBL旁系同源物之间的交叉反应性的方法。这些试剂使我们能够定义许多单个配音的可能特异性，从而得出了关于不同类别的DUB表现出的活动范围的一些令人惊讶的结论，并提出了开发和表征抑制剂的新机会，这些机会特异性地干扰了UBL蛋白质代谢的某些途径。在接下来的时期：我们将在定量标记和动力学方法中使用UBL-X试剂面板来确定来自对照和压力细胞中裂解物中活性去果糖酶的量和旁系同源性特异性。将特异性与纯化的催化域的特异性进行比较。我们将检验以下假设：响应细胞应激的sumoylated蛋白的细胞水平的变化受到活性去氨酰基酶的水平控制。我们将使用UBL-X试剂面板来确定SARS病毒类蛋白酶样蛋白酶表现出的配音活性的酶学特异性。该信息将提示可能的细胞底物，并用于安装高吞吐药物筛查，以鉴定可能在限制冠状病毒感染传播的抗病毒剂。我们将使用一组UBL-X试剂来评估据报道抑制去泛素化酶的抑制剂的特异性。这将提供一种快速而有选择性的方法，以评估“特定”抑制剂对去泛素化酶谱的全局影响。从广义上讲，这些研究将有助于我们对UBL蛋白质代谢的理解，并指导抑制剂和药物的开发，以治疗癌症和微生物感染。