Catalytic and regulatory mechanisms of human Tryptophan Dioxygenase

人色氨酸双加氧酶的催化和调节机制

• 批准号: 9107183
• 负责人: Syun-Ru Yeh
• 金额: $ 40.92万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE, INC
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9107183
• 关键词: Achievement; Active Sites; Alzheimer' s Disease; Amines; Binding; Biology; Catalysis; Chemistry; Collaborations; Crystallography; Degradation Pathway; Development; Dioxygen; Dioxygenases; Documentation; Ensure; Enzymes; Foundations; Freezing; Goals; Helix-Loop-Helix Motifs; Heme; Hemeproteins; Hepatic; Homeostasis; Human; Huntington Disease; Immunity; Iron; Kinetics; Knowledge; Kynurenine; Link; Liver; Malignant Neoplasms; Mental Depression; Methods; Microscopic; Molecular; Molecular Conformation; Mutagenesis; Neurodegenerative Disorders; Occupations; Optics; Pathway interactions; Peptides; Play; Positioning Attribute; Preparation; Property; Proteins; Protocols documentation; Public Health; Reaction; Regulation; Research; Role; Spectrum Analysis; Structure; Testing; Therapeutic; Therapeutic Intervention; Tryptophan; Ubiquitin; Ubiquitination; Water; analog; base; biophysical analysis; design; enzyme structure; flexibility; inhibitor/antagonist; insight; novel; public health relevance; simulation; therapeutic target; three dimensional structure; tumor; tumor growth

 DESCRIPTION (provided by applicant): Human tryptophan dioxygenase (hTDO) is an important hemeprotein that catalyzes the initial and rate-limiting step of the kynurenine (KYN) pathway-the conversion of Trp to N-formyl kynurenine (NFK). It is well-accepted that hTDO plays a critical role in controlling Trp homeostasis in liver. More recently, it was discovered tha hTDO is also expressed in a variety of human tumors to suppress antitumor immunity, thereby promoting aggressive tumor growth. In addition to cancer, hTDO has been implicated in a wide spectrum of neurodegenerative diseases, such as Alzheimer's and Huntington's diseases, as well as depression. hTDO has hence been recognized as an important therapeutic target. Despite its importance, the progress has been limited by the poor knowledge of the molecular properties of the enzyme. Recently, we have made several major breakthroughs in (i) developing a new protocol allowing the preparation of high quality hTDO protein, allowing the documentation of the first glimpse of the structural features critical for the dioxygenase chemistry, as well as an unusual activity of the ferric enzyme, (ii) solving the first substrate-bound 3D-structure of hTDO, as well as trapping reaction intermediates in single crystals and solved their structures, which unveiled several previously unidentified structural features that ar critical for controlling the catalytic activity and cellular degradation of the enzyme, one of the most rapidly degraded enzymes in liver, and (iii) obtaining the first evidence supporting the cellular degradation of hTDO via a ubiquitination-linked proteasomal degradation pathway. These important achievements allow us to build novel hypotheses and propose a comprehensive approach from a unique perspective to test them. The main objective of this project is to delineate the structure, function and regulation of hTDO, as part of our long term goal in comprehending heme dioxygenase chemistry and TDO biology. The specific aims of our project are: (i) identify the novel dioxygenase intermediates of hTDO. (ii) Determine the dioxygen activation mechanism of the unique ferric reaction. (iii) Delineate the product release mechanism of hTDO. (iv) Delineate the exosite-dependent regulation mechanisms of hTDO. We anticipate that fulfilling the objectives of this project will result in new insights into the struture, function and regulation of this essential enzyme, and provide an important blueprint for the design of hTDO-selective inhibitors as therapeutics for cancer, neurodegenerative diseases and depression.

 描述（申请人提供）：人色氨酸双加氧酶（hTDO）是一种重要的血红素蛋白，其催化犬尿氨酸（KYN）途径的起始和限速步骤——Trp向N-甲酰犬尿氨酸（NFK）的转化。 - 公认 hTDO 在控制肝脏色氨酸稳态方面发挥着关键作用。最近，人们发现 hTDO 具有重要作用。 hTDO 还在多种人类肿瘤中表达，可抑制抗肿瘤免疫，有效促进肿瘤生长。 除了癌症之外，hTDO 还与多种神经退行性疾病有关，例如阿尔茨海默病、亨廷顿病以及抑郁症。因此，尽管它很重要，但由于对酶的分子特性了解不足，进展受到限制。最近，我们在（i）开发允许制备的新方案方面取得了几项重大突破。高品质hTDO 蛋白，允许记录对双加氧酶化学至关重要的结构特征的第一眼，以及三价铁酶的不寻常活性，(ii) 解决 hTDO 的第一个底物结合 3D 结构，以及捕获单晶中的中间体并解析了它们的结构，揭示了一些以前未识别的结构特征，这些特征对于控制酶的催化反应和细胞降解至关重要，酶是肝脏中降解最快的酶之一，并且（iii）通过泛素化相关的蛋白酶体降解途径获得支持 hTDO 细胞降解的第一个证据。这些重要成就使我们能够建立新的假设，并从独特的角度提出一种全面的方法来测试它们。该项目的主要目标是描述。 hTDO 的结构、功能和调控，作为我们理解血红素双加氧酶化学和 TDO 生物学的长期目标的一部分，我们项目的具体目标是：(i) 识别新型双加氧酶中间体。 (ii) 确定独特的铁反应的双氧激活机制。 (iii) 描述 hTDO 的产物释放机制。 (iv) 描述 hTDO 的外位点依赖性调节机制。对这种必需酶的结构、功能和调节产生了新的见解，并为设计 hTDO 选择性抑制剂作为癌症、神经退行性疾病和抑郁症的治疗方法提供了重要的蓝图。