PURINE AND PYRIMIDINE METABOLISM

嘌呤和嘧啶代谢

• 批准号: 8361601
• 负责人: STEVEN E EALICK
• 金额: $ 1.14万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8361601
• 关键词: Adenosine; Adenosine Kinase; Allantoin; Anabolism; Animals; Anti-Bacterial Agents; Antibiotics; Aromatic Compounds; Asia; Bacteria; Biochemical; Biodegradation; Burkholderia; C-glycoside; Chemistry; Cleaved cell; Cytidine; Degradation Pathway; Deoxyribonucleosides; Dioxygen; Disease; Drug Delivery Systems; Energy Transfer; Enzymes; Escherichia coli; Exhibits; Flavin Mononucleotide; Funding; Gene Cluster; Genetic; Glycoside Hydrolases; Grant; Hydrolase; Hypoxanthines; Industrial fungicide; Infection; Isomerism; Klebsiella; Klebsiella pneumonia bacterium; Lyase; Malignant Neoplasms; Metabolic Pathway; Modification; Mono-S; National Center for Research Resources; Nitrogen; Nucleic Acids; Nucleosides; Nucleotides; Organism; Orotate Phosphoribosyltransferase; Orotidine-5' -Phosphate Decarboxylase; Oxygenases; Parasites; Parasitic infection; Pathway interactions; Phosphorylation; Phosphotransferases; Plants; Play; Principal Investigator; Production; Protein Biosynthesis; Proteins; Pseudouridine; Purine Nucleotides; Purine-Nucleoside Phosphorylase; Purines; Pyrimidine; Pyrimidine Nucleotides; RNA; Reaction; Research; Research Infrastructure; Resources; Ribonucleosides; Ribosomal RNA; Rice; Role; Ruta; Screening procedure; Signal Transduction; Small Nuclear RNA; Small Nucleolar RNA; Source; Structure; Substrate Specificity; Thymine; Toxic effect; Toxoplasmosis; Transfer RNA; Transgenic Organisms; United States; United States National Institutes of Health; Uracil; Uridine; Uridine Monophosphate; Uridine Phosphorylase; Work; base; combat; cost; inorganic phosphate; insight; interest; mildew; novel; plant fungi; purine; purine metabolism; purine/pyrimidine metabolism; ribose 1-phosphate; ribose-5-phosphate; structural biology; tool; transmethylation

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Pyrimidine and purine nucleotides are essential building blocks for the synthesis of nucleic acids and can also take part in energy transfer and storage, protein synthesis and signaling. Because of the importance of these molecules, the enzymes in their metabolic pathways represent potential drug targets for the treatment of many conditions including cancer and several types of parasitic infections. We have undertaken structural studies of various enzymes that play roles in the metabolism of pyrimidines and purines. Adenosine kinase (AK), a key enzyme in purine metabolism in parasites and a potential chemotherapeutic target for the treatment of Toxoplasma gondii infections, catalyzes the ATP dependent phosphorylation of adenosine. Purine nucleoside phosphorylase (PNP), which catalyzes the reversible phosphorolysis of ribonucleosides and 2'- deoxyribonucleosides to the free base and (2'-deoxy)ribose-1-phosphate, is an important enzyme for the salvage of purine nucleotides. RutA is a FMN dependent mono-oxygenase involved in a recently discovered pyrimidine degradation pathway that converts uracil (or thymine) to 3-hydroxypropionate (or 2-methyl-3-hydroxypropionate) in E. coli. Uridine phosphorylase (UP) catalyzes the reversible phosphorolysis of uridine with the formation of ribose-1-phosphate and uracil. Orotidine-5'-phosphate decarboxylase orotate phosphoribosyltransferase (OMPDC-OPRT) is a bifunctional enzyme that catalyzes the last two steps in the synthesis of uridine-5'-monophosphate (UMP). In addition to their many cellular uses, some organisms can metabolize nucleotides as a nitrogen source. Recent studies by two groups on Klebsiella sp. have revealed a gene cluster that is responsible for expressing the enzymes for utilizing purines as a sole nitrogen source in this organism. We have structurally characterized several of the enzymes that catalyze the breakdown of hypoxanthine to allantoin in Klebsiella pneumoniae in order to better understand this interesting pathway. Pseudouridine is the C-glycoside isomer of uridine and is the most abundant modification in RNA. It ubiquitously exists in tRNA, rRNA, snRNA and snoRNA. Recently, various biochemical, biophysical and genetic studies characterized two enzymes that are predominantly responsible for the biosynthesis of pseudouridine. Pseudouridine kinase (YeiC) phosphorylates pseudouridine to pseudouridine-5'-phosphate and pseudouridine glycosidase (YeiN) catalyzes the conversion from pseudouridine-5'-phosphate to uridine and ribose-5-phosphate by cleaving the C-C glycosidic bond. Structural studies of these enzymes will provide insights into the mechanism of pseudouridine biosynthesis and provide tools for the screening of possible antibacterial drugs that target this pathway. Mildiomycin is a peptidyl nucleoside antibiotic with strong activity against powdery mildew disease on plants and is used commercially as a fungicide. The initial steps of mildiomycin biosynthesis involve the cleavage of 5-hydroxymethyl cytidine-5'-monophosphate at the glycosidic bond by the enzyme MilB. It has been shown that MilB also catalyzes the cleavage reaction for cytidine-5'-monophosphate. The crystal structure of MilB was solved to better understand how its structure varies from other nucleotide hydrolases and how it provides its substrate specificity. Toxoflavin is an azapteridine that is poisonous to many plants, fungi, animals, and bacteria. Recently, toxoflavin has gained increasing interest because infection of rice plants by toxoflavin-producing bacteria such as Burkholderia glumae has led to a substantial loss of rice crops in the United States and Asia. Several of the steps in the biosynthesis of toxoflavin involve uncharacterized proteins that may potentially exhibit novel chemistry. Specifically, the proteins ToxC and/or ToxD appear to catalyze nitrogen-nitrogen bond formation which is poorly understood and the protein ToxA is predicted to catalyze sequential transmethylation reactions in the final step of toxoflavin production. To combat the toxicity of toxoflavin, recent work has exploited the ability of some enzymes to utilize dioxygen in the biosynthesis and biodegradation of numerous aliphatic and aromatic compounds. Recent work has led to the successful production of transgenic rice plants which express the putative oxygenase toxoflavin lyase (TflA) of Paenibacillus polymyxa JH2 to combat the deleterious effects of toxoflavin-producing bacteria on rice.

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 嘧啶和嘌呤核苷酸是合成核酸的重要构件，也可以参与能量转移和储存、蛋白质合成和信号转导。由于这些分子的重要性，它们代谢途径中的酶代表着治疗许多疾病的潜在药物靶点，包括癌症和几种寄生虫感染。 我们已经进行了各种酶的结构研究，这些酶在嘧啶和嘌呤的代谢中发挥作用。腺苷激酶(AK)是寄生虫体内嘌呤代谢的关键酶，也是治疗弓形虫感染的潜在化疗靶点，它催化ATP依赖的腺苷磷酸化。嘌呤核苷磷酸化酶(PNP)催化核糖核苷和2‘-脱氧核糖核苷可逆地磷酸化为游离碱和(2’-脱氧)核糖-1-磷酸，是回收嘌呤核苷酸的重要酶。RutA是一种依赖于FMN的单加氧酶，参与了新近发现的在大肠杆菌中将尿嘧啶(或胸腺嘧啶)转化为3-羟基丙酸酯(或2-甲基-3-羟基丙酸酯)的嘧啶降解途径。尿苷磷酸化酶(UP)催化尿苷的可逆磷酸化，生成核糖-1-磷酸和尿嘧啶。Orotidine-5‘-磷酸脱羧酶-Orotate磷酸核糖转移酶(OMPDC-OPRT)是催化尿苷-5’-单磷酸(UMP)合成最后两步的双功能酶。 除了它们在细胞上的许多用途外，一些生物还可以将核苷酸作为氮源进行代谢。最近两个小组对克雷伯氏菌的研究。已经揭示了一个基因簇，它负责表达利用嘌呤作为这种有机体中唯一氮源的酶。为了更好地理解这一有趣的途径，我们对几种催化肺炎克雷伯菌中次黄嘌呤分解为尿囊素的酶进行了结构表征。 假尿苷是尿苷的C-糖苷异构体，是RNA中含量最丰富的修饰。它普遍存在于tRNA、rRNA、SnRNA和snoRNA中。最近，各种生化、生物物理和遗传学研究确定了两种酶的特征，这两种酶主要负责假尿苷的生物合成。假尿苷酶(YEIN)通过裂解C-C糖苷键，催化假尿苷-5‘-磷酸转化为尿苷和核糖-5-磷酸。对这些酶的结构研究将有助于深入了解伪尿嘧啶生物合成的机制，并为筛选针对这一途径的可能的抗菌药物提供工具。 米迪霉素是一种多肽核苷类抗生素，对植物上的白粉病具有很强的活性，是一种商品化的杀菌剂。米迪霉素生物合成的初始步骤包括被MiLB酶在糖苷键上裂解5-羟甲基胞苷-5‘-单磷酸。结果表明，MiLB还催化了胞苷-5‘-单磷酸的裂解反应。为了更好地理解它的结构与其他核苷酸水解酶的不同，以及它如何提供底物专一性，我们解决了MiLB的晶体结构问题。 毒黄素是一种氮杂蝶啶，对许多植物、真菌、动物和细菌都有毒性。最近，毒黄素引起了人们越来越多的兴趣，因为产生毒黄素的细菌如Burkholderia glumae感染水稻植株，导致美国和亚洲的水稻作物大量损失。毒黄素生物合成的几个步骤涉及可能具有新化学特性的未知蛋白质。具体地说，ToxC和/或ToxD蛋白似乎催化氮-氮键的形成，而ToxA蛋白被预测在毒黄素生产的最后一步催化顺序的甲基化反应。为了对抗毒黄素的毒性，最近的工作利用了一些酶在许多脂肪族和芳香族化合物的生物合成和生物降解中利用氧气的能力。最近的工作已经成功地获得了表达多粘拟芽孢杆菌JH2加氧酶毒素黄素裂解酶(TflA)的转基因水稻植株，以对抗毒素产生细菌对水稻的有害影响。