Protein and small-molecule tools to probe the conformational dependence of the VCP/p97 protein-protein interaction network
用于探测 VCP/p97 蛋白质-蛋白质相互作用网络构象依赖性的蛋白质和小分子工具
基本信息
- 批准号:10228038
- 负责人:
- 金额:$ 34.07万
- 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:2018-09-05 至 2023-07-31
- 项目状态:已结题
- 来源:
- 关键词:ATP HydrolysisATP phosphohydrolaseAdaptor Signaling ProteinAddressAdoptedAffectAffinityAutophagosomeBacteriophagesBindingBiochemicalBiological AssayCell DeathCell physiologyCellsChemicalsComplexCouplingDegenerative DisorderDependenceDiseaseDisulfidesEngineeringEnzymesEquilibriumFamilyFunctional disorderFutureGoalsGolgi ApparatusHealthHomeostasisIndividualKnowledgeLeadLearningLibrariesLinkMalignant NeoplasmsMeasurementMeasuresMediatingMembraneMembrane ProteinsMolecularMolecular ConformationMutationN DomainNerve DegenerationNucleotidesOrangesOrganellesOutcomePathway interactionsPhage DisplayPhenotypePoint MutationPositioning AttributeProductionProtein ConformationProtein DynamicsProtein FamilyProtein InhibitionProteinsProteomeProteomicsReporterSiteStructureTestingTransfectionUbiquitinVariantWorkage relatedbasecancer cellcancer therapycell typedesigneffective therapyimprovedinhibitor/antagonistlink proteinmembermultisystem proteinopathymutantprotein complexprotein functionprotein protein interactionproteostasisscreeningsegregationsmall moleculetargeted treatmenttoolvalosin-containing protein
项目摘要
PROJECT SUMMARY/ABSTRACT
Modulating the functions of key players in protein homeostasis (proteostasis) could lead to therapies for diseases
from cancer to neurodegeneration. Valosin Containing Protein (VCP, p97), a member of the AAA+ (ATPases
associated with various cellular activities) family of enzymes, is one of the cell’s central regulators of proteostasis.
Functions as diverse as unfolding of ubiquitinated proteins from organelles, segregation of ubiquitinated proteins
from protein complexes, and remodeling of organelle membranes have been ascribed to VCP. These functions
are coordinated by a set of “adaptor” proteins and ubiquitin-processing enzymes that bind to VCP. Despite the
importance of these protein-protein interactions (PPI) to regulated proteostasis, there are major gaps in our
understanding of how adaptor proteins link VCP’s ATPase activity to diverse cellular functions. Furthermore,
point mutations in VCP cause a fatal, degenerative disease called Multisystem Proteinopathy 1 (MSP1). MSP1
is associated with multiple alterations in proteostasis that include both increased degradation of some proteins
and loss of degradation of others. VCP undergoes large conformational changes during ATP hydrolysis, and
MSP1 mutations alter VCPs conformational propensity. We and others have shown that PPI are also linked to
VCP conformation, leading to the hypothesis that VCP’s PPI network is modulated by ATPase-dependent
conformational dynamics, and that MSP1 mutations lead to dysregulation of the PPI network by altering these
dynamics. MSP1 disease should therefore be viewed as a disease of network dysregulation. To address this
hypothesis, we need new tools that address how adaptors bind to VCP conformations and alter ATPase activity,
and how conformational-dependent binding affects the cellular activities of the VCP network. We will address
these gaps through three Specific Aims. 1) We have shown that adaptor proteins sense VCP conformation.
We will extend these observations to at least eight adaptor/VCP complexes and will also evaluate the effect of
adaptors on ATPase activity and conformation. This analysis will provide predictions for which adaptor-
dependent functions are increased or inhibited in MSP1 cells. 2) We have utilized a site-directed small-molecule
discovery approach called disulfide-trapping to identify compounds that lock VCP into specific conformations.
We hypothesize that inhibiting VCP dynamics will stabilize some PPI, but will inhibit biochemical and cellular
functions that rely on VCP conformational dynamics. 3) We have developed phage-displayed libraries of the N-
domain of VCP to select mutants that bind with high affinity and selectivity to single adaptor proteins. We
hypothesize that blocking individual adaptor/VCP complexes in cells will lead to changes in VCP-mediated
pathways and the ubiquitin proteome (ubiquitinome). Combining PPI measurements, small-molecule
conformational locks, and protein-based PPI inhibitors will allow us to predict which VCP pathways lead to (or
mitigate) MSP1 phenotypes, and which should be harnessed to develop cancer-specific VCP inhibitors.
项目总结/摘要
调节蛋白质稳态(蛋白质稳态)中关键参与者的功能可能会导致疾病的治疗
从癌症到神经退化Valosin Containing Protein(VCP,p97)是AAA+(ATP酶)的成员
与各种细胞活动相关)酶家族,是细胞蛋白质稳态的中心调节因子之一。
功能多样,如从细胞器中展开泛素化蛋白,分离泛素化蛋白
从蛋白质复合物,和细胞器膜的重塑已被归因于VCP。这些功能
由一组“衔接”蛋白和结合VCP的泛素加工酶协调。尽管
这些蛋白质-蛋白质相互作用(PPI)的重要性,以调节蛋白质稳态,有很大的差距,在我们的
了解衔接蛋白如何将VCP的ATP酶活性与多种细胞功能联系起来。此外,委员会认为,
VCP中的点突变导致称为多系统蛋白质病1(MSP 1)的致命的退行性疾病。MSP1
与蛋白质稳态的多种改变有关,包括一些蛋白质降解增加
以及其他人的堕落。VCP在ATP水解过程中经历大的构象变化,
MSP 1突变改变VCP的构象倾向。我们和其他人已经表明,PPI也与
VCP构象,导致假设VCP的PPI网络是由ATP酶依赖的
MSP 1突变通过改变这些构象动力学,导致PPI网络的失调,
动力学因此,MSP 1疾病应被视为网络失调的疾病。为了解决这个
假设,我们需要新的工具来解决衔接子如何结合VCP构象并改变ATP酶活性,
以及构象依赖性结合如何影响VCP网络的细胞活性。我们将解决
这些差距通过三个具体目标。1)我们已经表明,衔接蛋白的意义VCP构象。
我们将这些观察扩展到至少八个适配器/VCP复合物,并将评估
衔接子对ATP酶活性和构象的影响。该分析将提供预测哪个适配器-
在MSP 1细胞中依赖性功能增加或抑制。2)我们利用定点小分子
一种称为二硫化物捕获的发现方法,用于识别将VCP锁定为特定构象的化合物。
我们假设,抑制VCP动力学将稳定一些PPI,但将抑制生化和细胞
依赖于VCP构象动力学的功能。3)我们已经开发了N-
结构域的VCP选择突变体,结合具有高亲和力和选择性的单一衔接蛋白。我们
假设阻断细胞中的单个接头/VCP复合物将导致VCP介导的
泛素蛋白质组(ubiquitinome)。结合PPI测量,小分子
构象锁,和蛋白质为基础的PPI抑制剂将使我们能够预测哪些VCP途径导致(或
缓解)MSP 1表型,并应利用其开发癌症特异性VCP抑制剂。
项目成果
期刊论文数量(6)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Modulating protein-protein interaction networks in protein homeostasis.
- DOI:10.1016/j.cbpa.2019.02.012
- 发表时间:2019-06
- 期刊:
- 影响因子:7.8
- 作者:Mengqi Zhong;Gregory M Lee;E. Sijbesma;C. Ottmann;M. Arkin
- 通讯作者:Mengqi Zhong;Gregory M Lee;E. Sijbesma;C. Ottmann;M. Arkin
Autophagy Receptor-Inspired Antibody-Fusion Proteins for Targeted Intracellular Degradation.
- DOI:10.1021/jacs.3c05199
- 发表时间:2023-11-08
- 期刊:
- 影响因子:15
- 作者:Jiang Z;Kuo YH;Arkin MR
- 通讯作者:Arkin MR
Multiparametric High-Content Assays to Measure Cell Health and Oxidative Damage as a Model for Drug-Induced Liver Injury.
- DOI:10.1002/cpch.90
- 发表时间:2020-12
- 期刊:
- 影响因子:0
- 作者:Pohan G;Espinosa JA;Chen S;Ang KK;Arkin MR;Markossian S
- 通讯作者:Markossian S
Synthetic autophagy receptor.
合成自噬受体。
- DOI:10.1080/15548627.2023.2278954
- 发表时间:2024
- 期刊:
- 影响因子:13.3
- 作者:Jiang,Ziwen;Kuo,Yu-Hsuan;Arkin,MichelleR
- 通讯作者:Arkin,MichelleR
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{{ truncateString('Michelle Arkin', 18)}}的其他基金
Development of caspase-6 inhibitors for treatment of NASH
开发治疗 NASH 的 caspase-6 抑制剂
- 批准号:
10608905 - 财政年份:2023
- 资助金额:
$ 34.07万 - 项目类别:
Optimizing brain penetrance of caspase-6 inhibitors to treat neurodegenerative diseases
优化 caspase-6 抑制剂的脑外显率来治疗神经退行性疾病
- 批准号:
10603619 - 财政年份:2023
- 资助金额:
$ 34.07万 - 项目类别:
Systematic stabilization of specific protein-protein interactions
特定蛋白质-蛋白质相互作用的系统稳定
- 批准号:
10703416 - 财政年份:2022
- 资助金额:
$ 34.07万 - 项目类别:
Systematic stabilization of specific protein-protein interactions
特定蛋白质-蛋白质相互作用的系统稳定
- 批准号:
10502096 - 财政年份:2022
- 资助金额:
$ 34.07万 - 项目类别:
Discovery of Small Molecule Ligands for PHD1 Reader Domain of Histone Demethylase KDM5A
组蛋白去甲基化酶 KDM5A 的 PHD1 阅读器结构域小分子配体的发现
- 批准号:
10183207 - 财政年份:2020
- 资助金额:
$ 34.07万 - 项目类别:
Discovery of Small Molecule Ligands for PHD1 Reader Domain of Histone Demethylase KDM5A
组蛋白去甲基化酶 KDM5A 的 PHD1 阅读器结构域小分子配体的发现
- 批准号:
10650159 - 财政年份:2020
- 资助金额:
$ 34.07万 - 项目类别:
Discovery of Small Molecule Ligands for PHD1 Reader Domain of Histone Demethylase KDM5A
组蛋白去甲基化酶 KDM5A 的 PHD1 阅读器结构域小分子配体的发现
- 批准号:
10442482 - 财政年份:2020
- 资助金额:
$ 34.07万 - 项目类别:
A high content screen dissecting ciliogenesis and oncogenic Hedgehog signaling
高内涵屏幕剖析纤毛发生和致癌 Hedgehog 信号传导
- 批准号:
9248278 - 财政年份:2016
- 资助金额:
$ 34.07万 - 项目类别:














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