Structural studies of DNA resolvases involved in hairpin telomere maintenance

参与发夹端粒维持的 DNA 解离酶的结构研究

• 批准号: 8513361
• 负责人: Hideki Aihara
• 金额: $ 27.02万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8513361
• 关键词: Address; Affinity; Agrobacterium; Bacteria; Bacteriophages; Binding; Biochemical; Biotechnology; Borrelia; Borrelia burgdorferi; Chemicals; Chemistry; Chromosomes; Cleaved cell; Complex; Cruciform DNA; DNA; DNA Fingerprinting; DNA Repair; DNA Sequence; DNA Structure; DNA biosynthesis; DNA strand break; DNA-Directed DNA Polymerase; Drug Design; Enzymes; Family; Food Industry; Fowlpox virus; Gene Rearrangement; Genetic; Genetic Recombination; Genome; HIV Integrase; Head; Holliday Junction Resolvases; Integrase; Length; Link; Lyme Disease; Maintenance; Mediating; Mitochondria; Molecular; Molecular Conformation; Natural regeneration; Nature; Nucleic Acids; Pathway interactions; Phosphotyrosine; Plants; Poxviridae; Process; Proteins; Reaction; Relapsing Fever; Research; Resolution; Resolvase; Role; Sequence Homology; Series; Site; Specificity; Structure; System; Tail; Telomere Maintenance; Virus; Work; chemical reaction; dimer; ds-DNA; endonuclease; human disease; member; pathogen; seal; telomere; tool development

DESCRIPTION (provided by applicant): Telomeres protect the ends of a linear chromosome and facilitate complete replication of terminal DNA sequences. The simplest form of telomere is a covalently closed hairpin structure found in bacteria and viruses carrying linear chromosomes, including members of the genus Borrelia - the causative agents of Lyme disease and relapsing fever - and the poxviruses. Replication of a linear chromosome with hairpin telomeres proceeds through a two-step mechanism, in which DNA polymerases first produce a concatenated replication intermediate that is subsequently resolved into unit-length chromosomes. This proposal focuses on the bacterial and poxviral enzymes that resolve the concatemeric chromosome to regenerate hairpin telomeres. The two classes of DNA resolvases utilize distinct types of chemical reactions to process the inverted repeat DNA sequences separating multiple copies of chromosomes. Bacterial protelomerases resolve a palindromic duplex substrate into hairpin products using the phophotyrosine-mediated DNA cleavage-rejoining reaction. On the other hand, the poxvirus resolvase binds specifically to the Holliday junction structure formed by hairpin extrusion at a palindromic sequence and makes symmetrical strand cleavages across the junction point. Even though the chemical natures of the reactions catalyzed by the bacterial and poxviral DNA resolvases are well established, it is poorly understood how these proteins adapt the respective catalytic modules to resolve DNA at the sites of replicated telomeres. In this proposal we will specifically address the following questions by determining crystal structures of various resolvase- DNA complexes: How do the bacterial protelomerase enzymes facilitate efficient refolding of a duplex substrate into hairpin products using the intrinsically isoenergetic DNA cleavage-rejoining chemistry? How does the poxvirus resolvase achieve high specificity in recognizing the branched DNA structure and catalyzing concerted strand cleavages at the junction point? Despite carrying out reactions seemingly independent of each other at the biochemical level, the two types of DNA resolvases may share a similar strategy in making symmetrical DNA cleavages across the inverted repeat junction. Our structural work will highlight diverse strategies as well as potentially a general mechanism employed by enzymes involved in the maintenance of hairpin telomeres in the important pathogens. Furthermore, our research may contribute to better understanding of many DNA rearrangement machineries that share similar reaction chemistries with the DNA resolvases studied here.

描述（由申请人提供）：端粒保护线性染色体的末端，并促进终端DNA序列的完全复制。端粒的最简单形式是一种在细菌和携带线性染色体的病毒中发现的共价闭合发夹结构，包括伯罗利属的成员 - 莱姆病和复发性发烧的病因 - 和痘病毒。与发夹端粒的线性染色体的复制是通过两步机制进行的，其中DNA聚合酶首先产生一个串联的复制中间体，随后将其分解为单位长染色体。该提议着重于解决串联染色体以再生发夹端粒的细菌和鼻虫酶。两类的DNA分辨率利用不同类型的化学反应来处理分隔多个染色体副本的倒重复DNA序列。细菌原始的原始蛋白酶酶使用凤凰介导的DNA裂解反应反应来解决一部分双链体底物进入发夹产物中。另一方面，鳞病毒分辨率专门结合了由发夹挤出在palindromic序列上形成的霍利迪连接结构，并使对称的链裂解在整个接线点上。即使通过细菌和蛇毒DNA分辨率催化的反应的化学生是良好的，但鲜为人知的这些蛋白如何适应各个催化模块以在复制端粒的位点解决DNA。在该提案中，我们将通过确定各种分解DNA复合物的晶体结构来特异性解决以下问题：细菌原始蛋白酶酶如何使用固有的等值异构剂DNA固定化的化学促进双链底物的有效重折叠到发夹产物中？在识别交界点处的分支DNA结构和催化协同的链裂解方面，鼠盆分解如何获得高特异性？尽管在生物化学水平上进行了似乎彼此独立的反应，但两种类型的DNA分辨率可能具有类似的策略，可以在整个反向重复连接中进行对称的DNA裂解。我们的结构性工作将突出不同的策略，并可能是参与重要病原体中发夹端粒涉及的酶采用的一般机制。此外，我们的研究可能有助于更好地理解许多DNA重排机器，这些机械与此处研究的DNA分辨率共享相似的反应化学。