DNA Helicases: Mechanisms and Functions
DNA 解旋酶:机制和功能
基本信息
- 批准号:8323299
- 负责人:
- 金额:$ 27.61万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2011
- 资助国家:美国
- 起止时间:2011-09-01 至 2015-08-31
- 项目状态:已结题
- 来源:
- 关键词:ATP HydrolysisAccountingActive SitesAddressAmino Acid MotifsBacteriophage T4BindingBinding ProteinsBinding SitesBiochemicalBiologicalBiological AssayBiological ModelsBiological ProcessChemicalsCouplingDNADNA BindingDNA FootprintDNA annealingDNA-Binding ProteinsDNA-Protein InteractionDataDefectDeuteriumDiseaseEnzymatic BiochemistryEnzymesExhibitsFamilyFundingGenomic InstabilityHereditary DiseaseHomologous GeneHuman GeneticsHydrogenIndividualInvestigationKineticsKnowledgeLeadLinkMalignant NeoplasmsMass Spectrum AnalysisMetabolismMethodsMolecularMutagenesisMutationPathway interactionsPlayPremature aging syndromeProcessProtein DynamicsProteinsRNAReactionReportingResearchResearch Project GrantsRoleSH3 DomainsStructureStructure-Activity RelationshipSurfaceTertiary Protein StructureTestingTimeWorkdimerenzyme coupling mechanismenzyme mechanismhelicasemeltingmembernovelprotein protein interactionprotein structurequadruplex DNArecombinational repairresearch studysingle moleculetoolunpublished works
项目摘要
DESCRIPTION (provided by applicant): Helicases are ubiquitous enzymes involved in virtually every aspect of DNA and RNA metabolism. This project focuses on one of the largest classes of this family of enzymes, superfamily 1B (SF1B). Limited structural information has slowed progress of our understanding of this class of enzymes. SF1B helicases couple ATP hydrolysis to DNA unwinding, but the rate limiting steps in this process are unknown. Specific amino acid motifs are known to make contact with DNA, but the dynamic role of these motifs has only been inferred. SF1B helicases interact with other proteins such as single-stranded binding proteins, but the biochemical and biological roles of these interactions are largely unaddressed. The importance of filling in these gaps in our knowledge relates to the many roles that helicases play in DNA metabolism including replication, repair, and recombination. Molecular defects in helicase activity have been directly linked to numerous human genetic diseases characterized by genome instability, premature aging, and cancer. Therefore, it is critical that we understand the mechanisms of these enzymes in order to understand how defects at the molecular level can lead to such devastating diseases. Dda helicase from bacteriophage T4 has served as the prototypical model system for the SF1B helicases. New structural data for Dda has led us to propose a mechano-chemical coupling mechanism that involves domains that include the standard helicase motifs along with novel domains that are uncharacterized. Helicase assays and DNA footprinting will be used to test this mechanism. We will determine the kinetic mechanism for ATP hydrolysis during DNA unwinding to determine the overall rate-limiting step in the process, which is currently unknown. Protein domains that are proposed to drive the helicase through conformational changes will be examined by rapid chemical footprinting methods that reveal whether DNA is bound tightly or loosely within the active site. High mobility protein motifs will be identified by hydrogen-deuterium exchange in order to determine the relationship between protein structure and dynamics. One of the major unanswered questions in helicase enzymology relates to the interaction between the enzyme and each individual strand of DNA. A combination of x-ray crystallographic, mass spectrometric and kinetic approaches will be used to identify all of the DNA binding sites on the surface of the enzyme. The structure-function relationship of these novel DNA binding sites will be determined through DNA unwinding experiments. The mechanism by which helicases remove proteins from DNA will be investigated using single molecule approaches. The role of protein-protein interactions will be determined by creating a tethered, dimeric form of the helicase and examining the ability of this enzyme to displace DNA-bound proteins. Answers to the questions posed in this proposal will advance the field in depth (helicase enzymology) and breadth (helicase interactions with protein partners), each of which will facilitate understanding of the role that these enzymes play in normal and pathogenic pathways of DNA metabolism. This work will provide experimental and conceptual tools to investigate other classes of helicases.
描述(由申请人提供):解旋酶是普遍存在的酶,几乎涉及 DNA 和 RNA 代谢的各个方面。该项目重点关注该酶家族中最大的一类,即超家族 1B (SF1B)。有限的结构信息减缓了我们对此类酶的理解进展。 SF1B 解旋酶将 ATP 水解与 DNA 解旋结合起来,但此过程中的限速步骤尚不清楚。已知特定的氨基酸基序会与 DNA 接触,但这些基序的动态作用仅是推测的。 SF1B 解旋酶与其他蛋白质(例如单链结合蛋白)相互作用,但这些相互作用的生化和生物学作用在很大程度上尚未得到解决。填补我们知识空白的重要性与解旋酶在 DNA 代谢中发挥的许多作用有关,包括复制、修复和重组。解旋酶活性的分子缺陷与许多以基因组不稳定、过早衰老和癌症为特征的人类遗传疾病直接相关。因此,我们必须了解这些酶的机制,以便了解分子水平的缺陷如何导致此类毁灭性疾病。来自噬菌体 T4 的 Dda 解旋酶已作为 SF1B 解旋酶的原型模型系统。 Dda 的新结构数据使我们提出了一种机械化学耦合机制,该机制涉及包含标准解旋酶基序的结构域以及未表征的新结构域。解旋酶测定和 DNA 足迹将用于测试这一机制。我们将确定 DNA 解旋过程中 ATP 水解的动力学机制,以确定该过程中的总体限速步骤,目前尚不清楚。通过构象变化驱动解旋酶的蛋白质结构域将通过快速化学足迹方法进行检查,揭示 DNA 在活性位点内是紧密结合还是松散结合。高迁移率蛋白质基序将通过氢-氘交换来鉴定,以确定蛋白质结构和动力学之间的关系。解旋酶学中尚未解答的主要问题之一涉及酶与每条 DNA 链之间的相互作用。 X 射线晶体学、质谱和动力学方法的结合将用于识别酶表面上的所有 DNA 结合位点。这些新的DNA结合位点的结构-功能关系将通过DNA解旋实验来确定。将使用单分子方法研究解旋酶从 DNA 中去除蛋白质的机制。蛋白质-蛋白质相互作用的作用将通过创建解旋酶的束缚二聚体形式并检查该酶取代 DNA 结合蛋白质的能力来确定。对本提案中提出的问题的回答将推动该领域的深度(解旋酶酶学)和广度(解旋酶与蛋白质伙伴的相互作用),每一个都将有助于理解这些酶在 DNA 代谢的正常和致病途径中所发挥的作用。这项工作将为研究其他类别的解旋酶提供实验和概念工具。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
数据更新时间:{{ journalArticles.updateTime }}
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
Kevin Douglas Raney其他文献
Kevin Douglas Raney的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('Kevin Douglas Raney', 18)}}的其他基金
Functions and Mechanisms of Helicases and G-Quadruplex Nucleic Acids
解旋酶和 G-四链体核酸的功能和机制
- 批准号:
9277158 - 财政年份:2017
- 资助金额:
$ 27.61万 - 项目类别:
Functions and Mechanisms of Helicases and G-Quadruplex Nucleic Acids
解旋酶和 G-四链体核酸的功能和机制
- 批准号:
9892786 - 财政年份:2017
- 资助金额:
$ 27.61万 - 项目类别:
Functions and Mechanisms of Helicases and G-Quadruplex Nucleic Acids
解旋酶和 G-四链体核酸的功能和机制
- 批准号:
9912771 - 财政年份:2017
- 资助金额:
$ 27.61万 - 项目类别:
G-quadruplex DNA as a chemical signaling agent
G-四链体 DNA 作为化学信号剂
- 批准号:
9010374 - 财政年份:2015
- 资助金额:
$ 27.61万 - 项目类别:
HCV NS3 and NS5A: Biochemical Mechanisms and Biological Functions
HCV NS3 和 NS5A:生化机制和生物学功能
- 批准号:
7842164 - 财政年份:2009
- 资助金额:
$ 27.61万 - 项目类别:
相似海外基金
Unraveling the Dynamics of International Accounting: Exploring the Impact of IFRS Adoption on Firms' Financial Reporting and Business Strategies
揭示国际会计的动态:探索采用 IFRS 对公司财务报告和业务战略的影响
- 批准号:
24K16488 - 财政年份:2024
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Early-Career Scientists
Mighty Accounting - Accountancy Automation for 1-person limited companies.
Mighty Accounting - 1 人有限公司的会计自动化。
- 批准号:
10100360 - 财政年份:2024
- 资助金额:
$ 27.61万 - 项目类别:
Collaborative R&D
Accounting for the Fall of Silver? Western exchange banking practice, 1870-1910
白银下跌的原因是什么?
- 批准号:
24K04974 - 财政年份:2024
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
CPS: Medium: Making Every Drop Count: Accounting for Spatiotemporal Variability of Water Needs for Proactive Scheduling of Variable Rate Irrigation Systems
CPS:中:让每一滴水都发挥作用:考虑用水需求的时空变化,主动调度可变速率灌溉系统
- 批准号:
2312319 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Standard Grant
A New Direction in Accounting Education for IT Human Resources
IT人力资源会计教育的新方向
- 批准号:
23K01686 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
An empirical and theoretical study of the double-accounting system in 19th-century American and British public utility companies
19世纪美国和英国公用事业公司双重会计制度的实证和理论研究
- 批准号:
23K01692 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
An Empirical Analysis of the Value Effect: An Accounting Viewpoint
价值效应的实证分析:会计观点
- 批准号:
23K01695 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Accounting model for improving performance on the health and productivity management
提高健康和生产力管理绩效的会计模型
- 批准号:
23K01713 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
New Role of Not-for-Profit Entities and Their Accounting Standards to Be Unified
非营利实体的新角色及其会计准则将统一
- 批准号:
23K01715 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Improving Age- and Cause-Specific Under-Five Mortality Rates (ACSU5MR) by Systematically Accounting Measurement Errors to Inform Child Survival Decision Making in Low Income Countries
通过系统地核算测量误差来改善特定年龄和特定原因的五岁以下死亡率 (ACSU5MR),为低收入国家的儿童生存决策提供信息
- 批准号:
10585388 - 财政年份:2023
- 资助金额:
$ 27.61万 - 项目类别: