Structural basis of phosphoinositide biosynthesis in Mycobacterium tuberculosis

结核分枝杆菌中磷酸肌醇生物合成的结构基础

• 批准号: 8953854
• 负责人: Filippo Mancia
• 金额: $ 24万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8953854
• 关键词: Active Sites; Alcohols; Amino Acids; Anabolism; Antibiotic Therapy; Archaea; Bacteria; Binding; Biological Assay; CDPdiacylglycerol-inositol 3-phosphatidyltransferase; Cell Wall; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chemicals; Complex; Computer Simulation; Coupled; Crystallization; Cytidine Diphosphate Diglycerides; Data; Development; Disadvantaged; Disease; Drug Design; Drug Targeting; Drug resistance; Engineering; Environment; Enzymes; Family; Generations; Genus Mycobacterium; Glycerophospholipids; Glycolipids; Goals; Immune response; In Vitro; Inflammatory Response; Inositol; Inositol Phosphates; Knowledge; Libraries; Life; Lipid Bilayers; Lipids; Mannose; Mannosides; Measures; Membrane; Modeling; Molecular; Monitor; Mutagenesis; Mycobacterium tuberculosis; Pathway interactions; Pharmaceutical Preparations; Phase; Phosphatidylinositol Phosphates; Phosphatidylinositols; Phospholipids; Phosphotransferases; Play; Population; Positioning Attribute; Process; Radiolabeled; Reaction; Reactive Oxygen Species; Recombinants; Research; Resolution; Route; Specificity; Staging; Structure; Substrate Specificity; Symptoms; Testing; Therapeutic; Time; Tuberculosis; X-Ray Crystallography; base; cell envelope; cost; design; disorder prevention; electron density; enzyme mechanism; enzyme structure; high throughput screening; improved; inhibitor/antagonist; inorganic phosphate; insight; lipoarabinomannan; lipomannan; macrophage; milligram; miniaturize; mycobacterial; novel; pathogen; public health relevance; radiotracer; research study; screening; virtual

 DESCRIPTION (provided by applicant): Tuberculosis (TB) is one of the world's deadliest diseases. Currently a third of the world's population is infected by Mycobacterium tuberculosis (MyTB), the causative agent for TB. In 2012 alone almost 9 million people developed TB resulting in 1.3 million deaths. Current antibiotic therapies have limited efficacy and must typically be used in combination and for prolonged periods of time. Furthermore, drug-resistant forms of MyTB are increasingly prevalent, underscoring the urgency to identify new targets and develop novel drugs against TB. The key determinants of all MyTB-host interactions are glycolipids in the cell wall of the pathogen that all share a common phosphatidylinositol (PI) anchor to the membrane and indeed PI is essential for MyTB survival. The crucial first step in biosynthesis of mycobacterial PI is carried out by an enzyme called phosphatidylinositolphosphate synthase (PIPS). PIPS belongs to a large family of enzymes called the CDP-alcohol phosphotransferases (CDP-AP). CDP-APs play the key role in the biosynthesis of all glycerophospholipids across all kingdoms of life. We have determined the atomic resolution structure by X-ray crystallography of the first representative CDP-AP and we now have solved a second structure of a PIPS which is 40% identical to the MyTB enzyme. Here we propose: (i) to use this PIPS model to study in molecular detail PI synthesis in MyTB (Aim 1); (ii) to build upon our expertise with CDP-APs to determine the structure of MyTB PIPS (Aim 2); (iii) to capitalize on our capability to generate abundant quantities of functional recombinant MyTB PIPS to measure its activity and set up a high-throughput screening assay for MyTB PIPS inhibitors whilst in parallel exploring chemical space by virtual screening to identify potential leads. Our research will not only offer unprecedented insight into how PI is made in MyTB, but it will also allow us to initiate structure-guided drug design efforts targeting the synthesis of this essential lipid. PIPS represents a key target for development of narrow spectrum anti- mycobacterial therapeutics. Indeed, the substrate for MyTB PIPS is unique to Archaea and few bacteria including Mycobacteria. In contrast, eukaryotic PI-synthases do not process inositol phosphate and most bacteria lack phosphatidylinositol entirely. We are in a unique position to set this framework given our results and expertise on the structures of CDP-APs and PIPS.

 描述（由应用程序提供）：结核病（TB）是世界上最致命的疾病之一。目前，世界人口的三分之一被结核分枝杆菌（MYTB）感染，TB的致病剂。仅在2012年，就有近900万人出现了结核病，导致130万人死亡。当前的抗生素疗法的有效性有限，通常必须用于组合和长时间。此外，MYTB的耐药形式越来越普遍，强调了识别新靶标的紧迫性并开发针对TB的新型药物。所有MYTB-host相互作用的关键决定剂是病原体细胞壁中的糖脂，它们都与膜上共同呈磷脂酰肌醇（PI）锚固，而PI的确对MYTB生存至关重要。分枝杆菌PI生物合成的关键第一步是通过一种称为磷脂酰辛迪糖磷酸合酶（PIPS）的酶进行的。 PIPS属于一个称为CDP钙磷酸转移酶（CDP-AP）的大型酶。 CDP-APS在所有王国的所有甘油磷脂的生物合成中都起着关键作用。我们已经通过第一个代表性CDP-AP的X射线晶体学确定了原子分辨率结构，现在我们解决了PIP的第二个结构，在这里我们提出的是40％相同：（i）使用此PIPS模型在MyTB中使用分子细节PI合成研究（AIM 1）； （ii）以CDP-APS来建立我们的专业知识，以确定MYTB PIP的结构（AIM 2）； （iii）利用我们的能力产生大量功能性重组MYTB PIP，以测量其活性并为MYTB PIPS抑制剂建立高通量筛选测定法，同时通过虚拟筛选以识别潜在的潜在潜在的潜在潜在客户，同时探索化学空间。我们的研究不仅将对MYTB中PI的制作方式提供前所未有的见解，而且还可以使我们能够发起针对合成这种基本脂质的结构引导的药物设计工作。 PIPS代表了狭窄频谱抗分枝杆菌疗法的发展的关键目标。实际上，MYTB PIPS的底物是古细菌和细菌很少的。包括分枝杆菌。相比之下，真核PI合成酶不会处理肌醇磷酸盐，大多数细菌完全缺乏磷脂酰肌醇。鉴于我们在CDP-APS和PIP的结构方面的结果和专业知识，我们处于独特的位置。