Purine Metabolism in Trichomonas vaginalis

阴道毛滴虫的嘌呤代谢

• 批准号: 7420991
• 负责人: Ching Chung WANG
• 金额: $ 28.18万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7420991
• 关键词: 2' -deoxyadenosine; 2-Fluoroadenine; 6-methylpurine; Adenine; Adenosine; Adenosine Kinase; Alcohols; Bacterial Typing; Binding; Biological Assay; Biological Availability; Catalysis; Cells; Chemicals; Cloning; Computer Graphics; Crystallography; Data; Depth; Disease; Disulfiram; Enzyme Inhibitor Drugs; Enzyme Inhibitors; Enzyme Kinetics; Enzymes; Escherichia coli; Future; Growth; Guanosine; HIV Infections; Human; Infection; Kinetics; Lead; Libraries; Medical; Metronidazole; Metronidazole resistance; Modeling; Modification; Monitor; Mutagenesis; Nucleosides; Nucleotides; Organism; Outcomes Research; Parasites; Pathway interactions; Pharmaceutical Preparations; Phosphorylase a; Play; Predisposition; Protein Sequence Analysis; Proteins; Purine Nucleoside Phosphorylase Inhibitor; Purine Nucleosides; Purine Nucleotides; Purine-Nucleoside Phosphorylase; Purines; Purpose; Pyrimidine Nucleotides; Reaction; Recombinant Proteins; Recombinants; Reporting; Research; Rodent; Role; Sexually Transmitted Diseases; Site-Directed Mutagenesis; Specificity; Structure; Structure-Activity Relationship; Substrate Specificity; Testing; Therapeutic; Toxic effect; Trichomonas Infections; Trichomonas vaginalis; United States; Urethritis; Vaginitis; Woman; X ray diffraction analysis; X-Ray Crystallography; X-Ray Diffraction; analog; base; chemical synthesis; chemotherapy; combinatorial; design; enzyme structure; enzyme substrate; formycin; guanosine kinase; improved; inhibitor/antagonist; kinase inhibitor; men; novel strategies; nucleoside kinase; pathogen; prototype; purine; purine metabolism; small molecule libraries; social

DESCRIPTION (provided by applicant): Trichomonas vaginalis is one of the most prevalent infectious pathogens with a worldwide distribution and about 5 million new cases occurring every year. It has vulnerability in lacking de novo synthesis of purine nucleotides and relying on the sequential actions of a purine nucleoside phosphorylase (PNP) and a purine nucleoside kinase (PNK) for salvaging the external purine bases. Inhibition of either enzyme can result in cessation of T. vaginalis growth and could thus lead to a therapeutic gain. T. vaginalis PNP is a bacterial type hexameric protein with an overwhelming catalytic efficiency in converting adenine to adenosine, whereas human PNP is a trimeric protein not even recognizing adenine as a substrate. T. vaginalis PNK is primarily a bacterial type guanosine kinase, whereas human PNK uses only adenosine as substrate. There are thus many opportunities for designing selective inhibitors against the two parasite enzymes. The parasite PNP was cloned, expressed and had its kinetic mechanism of catalysis well analyzed by us. In the future plan, its structure will be studied by crystallography and its structure-function relationship will be dissected by site-directed mutagenesis. Computer graphic modeling will be used for selecting specific enzyme inhibitors and combinatorial library synthesis will be involved in improving the lead compounds. Well known bacterial PNP inhibitors, such as formycin A, and the subversive substrates of Escherichia coli PNP will be tested and structurally modified for improved potency against the parasite enzyme. T. vaginalis PNK will be cloned sequenced and expressed. The kinetic mechanism of its catalytic function will be examined and compared with that of human adenosine kinase (AK). Following crystallographic structural analysis and site-directed mutagenesis to verify the mechanisms of substrate binding, product release, etc., computer graphic modeling and combinatorial library synthesis will be applied for selection of potent and specific enzyme inhibitors. Many well-known inhibitors of human AK with demonstrated bioavailability will be structurally modified to convert them from human AK inhibitors into potentially selective T. vaginalis PNK inhibitors for anti-Trichomoniasis chemotherapy. The prospect in a successful outcome from this research plan appears good.

描述（由申请人提供）：阴道毛滴虫是最流行的传染性病原体之一，在世界范围内分布，每年约有 500 万新病例发生。它的弱点在于缺乏嘌呤核苷酸的从头合成，并且依赖嘌呤核苷磷酸化酶（PNP）和嘌呤核苷激酶（PNK）的连续作用来挽救外部嘌呤碱基。抑制任何一种酶都可以导致阴道毛滴虫生长停止，从而获得治疗效果。阴道毛滴虫 PNP 是一种细菌型六聚蛋白，在将腺嘌呤转化为腺苷方面具有压倒性的催化效率，而人类 PNP 是一种三聚蛋白，甚至不识别腺嘌呤作为底物。阴道毛滴虫 PNK 主要是一种细菌型鸟苷激酶，而人类 PNK 仅使用腺苷作为底物。因此，有很多机会设计针对这两种寄生虫酶的选择性抑制剂。我们对寄生虫PNP进行了克隆、表达，并对其催化动力学机制进行了深入分析。未来的计划将通过晶体学研究其结构，并通过定点突变剖析其结构与功能关系。计算机图形模型将用于选择特定的酶抑制剂，组合库合成将涉及改进先导化合物。众所周知的细菌 PNP 抑制剂（例如福霉素 A）和大肠杆菌 PNP 的颠覆性底物将进行测试并进行结构修饰，以提高对抗寄生虫酶的效力。阴道毛滴虫 PNK 将被克隆、测序和表达。将检查其催化功能的动力学机制，并与人腺苷激酶（AK）进行比较。通过晶体结构分析和定点突变来验证底物结合、产物释放等机制，将应用计算机图形建模和组合文库合成来选择有效且特异性的酶抑制剂。许多众所周知的具有生物利用度的人类 AK 抑制剂将进行结构修饰，将其从人类 AK 抑制剂转化为潜在的选择性阴道毛滴虫 PNK 抑制剂，用于抗滴虫化疗。该研究计划取得成功的前景似乎良好。