Theoretical and experimental investigation of multi-domain protein folding and conformational dynamics
多域蛋白质折叠和构象动力学的理论和实验研究
基本信息
- 批准号:10218203
- 负责人:
- 金额:$ 45.81万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2018
- 资助国家:美国
- 起止时间:2018-09-01 至 2023-07-31
- 项目状态:已结题
- 来源:
- 关键词:AddressBindingBiological ProcessBiologyBypassCatalysisCell physiologyCellsChemicalsCircular DichroismClosure by clampCollaborationsCrowdingDNADNA BindingDNA DamageDNA lesionDNA-Directed DNA PolymeraseElementsEnergy TransferEnvironmentFamilyFluorescenceFluorescence SpectroscopyHot SpotHumanIn VitroInvestigationIonsKineticsLaboratoriesLaboratory ResearchLobeMeasurementMethodsMicroscopicModelingMolecularMolecular ConformationNamesNucleotidesOrganismPathway interactionsPlayPolymerasePrincipal InvestigatorProcessProliferating Cell Nuclear AntigenProtein ConformationProtein DynamicsProtein EngineeringProteinsPublishingResearch PersonnelRibosomesRoleSlideStructureSulfolobus solfataricusSystemTertiary Protein StructureTestingTheoretical StudiesTheoretical modelUncertaintyValidationWorkantigen bindingbaseconformational conversiondrug discoveryexperimental studyflexibilityimprovedin vivomacromoleculenovelpredictive modelingprotein foldingreplication factor Astopped-flow fluorescencethermostabilitythree dimensional structure
项目摘要
PROJECT SUMMARY
Proper folding is crucial to achieving a protein’s unique three dimensional structure while the conformational
dynamics of the protein play a major role in its biological function. Although significant progress has been
made in understanding the folding/unfolding and conformational dynamics for single-domain proteins, these
two fundamental processes remain largely unexplored for multi-domain proteins, which have been suggested
to account for up to 80% of all eukaryotic proteins. Therefore, a significant disparity exists in our understanding
of the underlying mechanisms of folding/unfolding and conformational dynamics for the majority of human
proteins and we seek to address these uncertainties through theoretical and experimental investigation. In this
proposal, we devise a comprehensive strategy to answer the above, in-depth mechanistic unknowns regarding
multi-domain proteins through an energy landscape approach with subsequent experimental validation. The
energy landscape approach significantly improves technical capabilities through the establishment of
theoretical models for uncovering underlying mechanisms. By establishing the microscopic energy landscape
and structure based models, we will elucidate the folding/unfolding mechanisms of DPO4, a multi-domain,
model Y-family DNA polymerase critical for bypassing unrepaired DNA lesions, in vitro and in vivo (here means
mimicking in vivo conditions), and predict possible intermediate states and critical residues under various
environments, including the presence of the ribosome (co-translational) and a crowding agent (in vivo), as well
as different thermal and chemical denaturant conditions. Through our microscopic energy landscape and
structure based models, we will reveal the underlying mechanisms of conformational changes between various
conformational states of DPO4 upon binding to DNA or a protein replication factor PCNA through quantifying
the stability, kinetics, and structural hot spots critical for function. The theoretical model predictions will be
tested and validated through stopped-flow, circular dichroism, fluorescence energy transfer, and other
spectroscopic experiments. The results generated from the proposal will advance the DNA polymerase field
while the methods developed here are general and can serve as a framework for studies of folding/unfolding
and conformational dynamics of other multi-domain proteins. Moreover, the intricacies of protein
folding/unfolding and conformational transitions revealed by our proposed studies will facilitate protein design
and drug discovery.
项目摘要
正确的折叠对于实现蛋白质独特的三维结构至关重要,而构象
蛋白质的动力学在其生物学功能中起主要作用。虽然取得了重大进展,
在理解折叠/展开和构象动力学的单结构域蛋白质,这些
对于多结构域蛋白质,有两个基本的过程仍然没有被探索,
占所有真核生物蛋白质的80%。因此,我们的理解存在很大的差异,
折叠/展开和构象动力学的基本机制,为大多数人类
蛋白质,我们试图通过理论和实验研究来解决这些不确定性。在这
建议,我们设计了一个全面的战略,以回答上述,深入的机械未知数,
多域蛋白质通过能量景观的方法与随后的实验验证。的
能源景观方法通过建立
揭示潜在机制的理论模型。通过建立微观能源景观
和基于结构的模型,我们将阐明DPO 4,一个多结构域,
模型Y-家族DNA聚合酶对于绕过未修复的DNA损伤至关重要,在体外和体内(这里指的是
模拟体内条件),并预测各种条件下可能的中间状态和关键残基。
环境,包括核糖体(共翻译)和拥挤剂(体内)的存在,以及
作为不同的热变性和化学变性条件。通过我们的微观能源景观,
基于结构的模型,我们将揭示各种构象变化之间的潜在机制,
通过定量测定DPO 4与DNA或蛋白质复制因子PCNA结合时的构象状态
稳定性、动力学和对功能至关重要的结构热点。理论模型预测将是
通过停流、圆二色性、荧光能量转移等测试和验证
光谱实验该提案产生的结果将推动DNA聚合酶领域的发展
虽然这里开发的方法是通用的,可以作为研究折叠/展开的框架,
和其他多结构域蛋白的构象动力学。此外,蛋白质的复杂性
我们的研究揭示的折叠/去折叠和构象转变将有助于蛋白质设计
和药物发现。
项目成果
期刊论文数量(8)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Investigating the Conformational Dynamics of a Y-Family DNA Polymerase during Its Folding and Binding to DNA and a Nucleotide.
- DOI:10.1021/jacsau.1c00368
- 发表时间:2022-02-28
- 期刊:
- 影响因子:8
- 作者:Chu X;Suo Z;Wang J
- 通讯作者:Wang J
Investigating the trade-off between folding and function in a multidomain Y-family DNA polymerase.
- DOI:10.7554/elife.60434
- 发表时间:2020-10-20
- 期刊:
- 影响因子:7.7
- 作者:Chu X;Suo Z;Wang J
- 通讯作者:Wang J
Kinetic Investigation of Translesion Synthesis across a 3-Nitrobenzanthrone-Derived DNA Lesion Catalyzed by Human DNA Polymerase Kappa
人类 DNA 聚合酶 Kappa 催化的 3-硝基苯并蒽酮衍生 DNA 损伤跨损伤合成的动力学研究
- DOI:10.1021/acs.chemrestox.9b00219
- 发表时间:2019
- 期刊:
- 影响因子:4.1
- 作者:Phi, Kenneth K.;Smith, Madison C.;Tokarsky, E. John;Suo, Zucai
- 通讯作者:Suo, Zucai
Conformational state switching and pathways of chromosome dynamics in cell cycle
- DOI:10.1063/5.0007316
- 发表时间:2020-09-01
- 期刊:
- 影响因子:15
- 作者:Chu, Xiakun;Wang, Jin
- 通讯作者:Wang, Jin
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{{ truncateString('JIN WANG', 18)}}的其他基金
Targeting the HIV-1 Reservoir in Myeloid Cells
靶向骨髓细胞中的 HIV-1 储存库
- 批准号:
10326730 - 财政年份:2021
- 资助金额:
$ 45.81万 - 项目类别:
Targeting the HIV-1 Reservoir in Myeloid Cells
靶向骨髓细胞中的 HIV-1 储存库
- 批准号:
10636954 - 财政年份:2021
- 资助金额:
$ 45.81万 - 项目类别:
Targeting the HIV-1 Reservoir in Myeloid Cells
靶向骨髓细胞中的 HIV-1 储存库
- 批准号:
10445309 - 财政年份:2021
- 资助金额:
$ 45.81万 - 项目类别:
Theoretical and experimental investigation of multi-domain protein folding and conformational dynamics
多域蛋白质折叠和构象动力学的理论和实验研究
- 批准号:
9769815 - 财政年份:2018
- 资助金额:
$ 45.81万 - 项目类别:
Molecular regulation of immunity to viral infections
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9028083 - 财政年份:2015
- 资助金额:
$ 45.81万 - 项目类别:
SELECTIVE MITOCHONDRIAL AUTOPHAGY IN THE MAINTENANCE OF GENOME STABILITY
选择性线粒体自噬维持基因组稳定性
- 批准号:
7963177 - 财政年份:2010
- 资助金额:
$ 45.81万 - 项目类别:
SELECTIVE MITOCHONDRIAL AUTOPHAGY IN THE MAINTENANCE OF GENOME STABILITY
选择性线粒体自噬维持基因组稳定性
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8089250 - 财政年份:2010
- 资助金额:
$ 45.81万 - 项目类别:
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