Structural Studies of Ribonucleotide Reductase Complexes

核糖核苷酸还原酶复合物的结构研究

• 批准号: 7806713
• 负责人: Nozomi Ando
• 金额: $ 4.76万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7806713
• 关键词: Active Sites; Affinity; Antineoplastic Agents; Binding; Biological Models; Chemistry; Complex; Crystallization; DNA biosynthesis; Data; Deoxyribonucleotides; Diphosphates; Docking; Drug Delivery Systems; Drug Design; Enzymes; Equilibrium; Escherichia coli; Future; Gases; Health; Human; Individual; Lasers; Methods; Modeling; Molecular; Molecular Sieve Chromatography; Molecular Weight; Organism; Oxidoreductase; Pharmaceutical Preparations; Phase; Proteins; Radial; Resolution; Ribonucleotide Reductase; Ribonucleotides; Roentgen Rays; Shapes; Site; Solutions; Structure; Time; X-Ray Crystallography; anticancer research; base; cancer therapy; design; dimer; gemcitabine; inhibitor/antagonist; insight; light scattering; monomer; protein structure; public health relevance; repaired; response

DESCRIPTION (provided by applicant): Ribonucleotide reductases (RNRs) are multi-subunit enzymes that are important targets for anti-cancer therapy and thus, directly relevant to human health. Intact complexes of class la RNRs, which include human and E. coli RNRs, have never been visualized by structural methods. Additionally, the oligomerization states of the inhibited class Ia RNR complexes are not well established, and it has been suggested that the inhibited RNRs may exist as higher order oligomers than the active complex. Incubation with the mechanism-based inhibitor and chemotherapeutic drug, gemcitabine (2',2'-difluoro-2'-deoxycytidine) diphosphate, has been shown to yield stable inhibited complexes of human and E. coli RNRs. Here, we propose a structural study of E. coli and human RNRs using X-ray crystallography and small-angle X-ray scattering (SAXS). Aim 1: To determine the quaternary structures of human and E. coli RNRs in solution using SAXS. The SAXS structures can be used to validate the docking model and future crystal structures of RNR complexes. The proposed solution SAXS studies will provide insight into how E. coli and human RNRs may form different oligomeric species in response to inhibitors and more significantly, how quaternary structure may regulate the overall activity of class la RNRs. Aim 2: To determine the crystal structure of human RNR inhibited by gemcitabine. Crystal structures of the gemcitabine-bound human RNR complex will allow us to identify how this drug may modify the active site and disrupt the catalytically essential radical transfer This structure will also allow us to observe the binding interactions between the subunits for the first time as well as provide insight into the future design of chemotherapeutic drugs that target this enzyme. PUBLIC HEALTH RELEVANCE: Ribonucleotide reductase is a protein found in all organisms that is an important target for cancer drugs such as gemcitabine. Unlike many other proteins that are drug targets, however, the complete structure of this protein has never been solved. By studying the structure of this protein, we will understand more about cancer drugs that are currently in use and aid future drug design efforts.

描述（由申请人提供）：核糖核苷酸还原酶（RNR）是多亚基酶，是抗癌治疗的重要靶标，因此与人类健康直接相关。 Ia类RNR的完整复合物，包括人类和大肠杆菌RNR，从未通过结构方法可视化。此外，受抑制的 Ia 类 RNR 复合物的寡聚状态尚未很好地确定，并且已经表明受抑制的 RNR 可能以比活性复合物更高阶的寡聚物存在。与基于机制的抑制剂和化疗药物吉西他滨（2',2'-二氟-2'-脱氧胞苷）二磷酸一起孵育已被证明可产生人和大肠杆菌 RNR 的稳定抑制复合物。在这里，我们提出使用 X 射线晶体学和小角 X 射线散射 (SAXS) 对大肠杆菌和人类 RNR 进行结构研究。目标 1：使用 SAXS 确定溶液中人类和大肠杆菌 RNR 的四级结构。 SAXS 结构可用于验证 RNR 复合物的对接模型和未来的晶体结构。所提出的解决方案 SAXS 研究将深入了解大肠杆菌和人类 RNR 如何响应抑制剂而形成不同的寡聚物种，更重要的是，四级结构如何调节 Ia 类 RNR 的整体活性。目标 2：确定吉西他滨抑制的人 RNR 的晶体结构。吉西他滨结合的人 RNR 复合物的晶体结构将使我们能够确定该药物如何修饰活性位点并破坏催化必需的自由基转移。这种结构也将使我们能够首次观察亚基之间的结合相互作用为针对这种酶的化疗药物的未来设计提供见解。 公共卫生相关性：核糖核苷酸还原酶是一种存在于所有生物体中的蛋白质，是吉西他滨等癌症药物的重要靶标。然而，与许多其他作为药物靶标的蛋白质不同，这种蛋白质的完整结构从未得到解决。通过研究这种蛋白质的结构，我们将更多地了解目前正在使用的抗癌药物，并有助于未来的药物设计工作。