Phosphorylation-dependent regulation of protein secretion in Mycobacterium tuberculosis
结核分枝杆菌蛋白质分泌的磷酸化依赖性调节
基本信息
- 批准号:10183156
- 负责人:
- 金额:$ 22.13万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2020
- 资助国家:美国
- 起止时间:2020-06-08 至 2023-05-31
- 项目状态:已结题
- 来源:
- 关键词:AddressAffectAllelesAnimal ModelArchaeaArginineBacteriaBiological AssayC-terminalCell physiologyCellsCo-ImmunoprecipitationsComplementCuesCytoplasmic TailDataDiseaseELF3 geneEnvironmentEscherichia coliFiltrationGene ExpressionGenesGeneticGenetic StructuresGoalsInfectionIntegral Membrane ProteinKnowledgeLaboratoriesLeadMediatingMembrane ProteinsMessenger RNAMethodsMycobacterium tuberculosisN-terminalOperonPathogenesisPathway interactionsPhosphoproteinsPhosphorylationPhosphorylation SitePhosphotransferasesPhysiologyPlayPost-Translational Protein ProcessingProcessProtein BiosynthesisProtein KinaseProtein SecretionProtein translocationProteinsProteomicsPublishingRegulationResearchRoleSignal TransductionStructureSystemTestingTranscriptional RegulationTuberculosisTwin Multiple BirthTwo-Hybrid System TechniquesWorkbaseenzyme activityexperimental studyhuman pathogeninsightnovelnovel therapeutic interventionpathogenprotein functionprotein protein interactionresponsesuccesstranscription factoryeast two hybrid system
项目摘要
PROJECT SUMMARY
The goal of this research is to investigate the role of phosphorylation of the extended N-terminal
domain of Mycobacterium tuberculosis SecE, an essential component of the SecYEG transmembrane
protein translocation channel, in regulating protein secretion via the general secretory pathway. The SecE
C-terminal domain encodes a transmembrane helix and an amphipathic helix that are highly conserved
across bacteria and archaea. While SecE in many bacteria comprises only the ~65 residues that encode
these features, others, such as E. coli and M. tuberculosis have an extended N-terminal domain. We have
identified three Ser or Thr residues in this domain that are phosphorylated, all of which show highly
significantly decreased phosphorylation in response to depletion of PknA and/or PknB, the two essential
Ser/Thr kinases of this pathogen. These findings provide the premises for our hypothesis that the SecE
extended N-terminal domain regulates protein secretion in a phosphorylation-dependent manner.
To investigate the regulatory role of this domain, we propose two Aims. In Aim 1 we will perform
quantitative proteomics of culture filtrates of PknA replete and PknA depleted M. tuberculosis strains. We
will also construct strains expressing native SecE or SecE with phosphomimetic or phosphoablative
substitutions at the identified phosphorylation sites, and quantify protein secretion in these strains. These
experiments will determine the effects of SecE N-terminal domain phosphorylation on protein secretion. In
Aim 2 we will identify proteins that interact with SecE using two complementary methods, i) co-
immunoprecipitation with SecE in M. tuberculosis and ii) two-hybrid assays to detect protein-protein
interactions with native, phosphomimetic and phosphoablative SecE. These experiments will suggest how
SecE and its phosphorylation may function in the general secretion pathway.
We anticipate that this research will reveal a novel mechanism for regulating protein secretion that will
be relevant for tuberculosis pathogenesis and may lead to new therapeutic approaches to treat this disease.
项目摘要
本研究的目的是探讨延长的N-末端磷酸化的作用,
结核分枝杆菌SecE的结构域,SecYEG跨膜的重要组成部分
蛋白质转运通道,通过一般分泌途径调节蛋白质分泌。SecE
C-末端结构域编码高度保守的跨膜螺旋和两亲性螺旋
细菌和古细菌之间。虽然许多细菌中的SecE仅包含约65个编码
这些特征,其他,如E. coli和M.结核病具有延伸的N-末端结构域。我们有
鉴定了该结构域中磷酸化的三个Ser或Thr残基,所有这些残基都显示出高度的磷酸化。
显著降低的磷酸化响应于PknA和/或PknB的消耗,这两种必需的磷酸化蛋白质是PknA和/或PknB的磷酸化蛋白质。
该病原体的Ser/Thr激酶。这些发现为我们假设SecE
延伸的N-末端结构域以磷酸化依赖性方式调节蛋白质分泌。
为了研究这个域的调节作用,我们提出了两个目标。在目标1中,
PknA充足和PknA缺失的M.结核菌株我们
还将构建表达天然SecE或具有磷酸模拟或磷酸消融的SecE的菌株
在鉴定的磷酸化位点处的取代,并定量这些菌株中的蛋白质分泌。这些
实验将确定SecE N-末端结构域磷酸化对蛋白分泌的影响。在
目的2我们将使用两种互补的方法鉴定与SecE相互作用的蛋白质,i)共-
用SecE免疫沉淀法检测M.结核病和ii)检测蛋白质-蛋白质的双杂交测定
与天然、磷酸模拟和磷酸消融SecE的相互作用。这些实验将揭示
SecE及其磷酸化可能在一般分泌途径中起作用。
我们预计这项研究将揭示一种调节蛋白质分泌的新机制,
与结核病发病机制有关,并可能导致治疗这种疾病的新的治疗方法。
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)
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ROBERT N HUSSON其他文献
ROBERT N HUSSON的其他文献
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{{ truncateString('ROBERT N HUSSON', 18)}}的其他基金
Regulation of VapC toxins by Ser/Thr phosphorylation of VapB antitoxins in M. tuberculosis
结核分枝杆菌中 VapB 抗毒素的 Ser/Thr 磷酸化对 VapC 毒素的调节
- 批准号:
9978276 - 财政年份:2020
- 资助金额:
$ 22.13万 - 项目类别:
Phosphorylation-dependent regulation of protein secretion in Mycobacterium tuberculosis
结核分枝杆菌蛋白质分泌的磷酸化依赖性调节
- 批准号:
10056045 - 财政年份:2020
- 资助金额:
$ 22.13万 - 项目类别:
Regulation of VapC toxins by Ser/Thr phosphorylation of VapB antitoxins in M. tuberculosis
结核分枝杆菌中 VapB 抗毒素的 Ser/Thr 磷酸化对 VapC 毒素的调节
- 批准号:
10112821 - 财政年份:2020
- 资助金额:
$ 22.13万 - 项目类别:
Development of a CRISPR interference system for mycobacteria
分枝杆菌 CRISPR 干扰系统的开发
- 批准号:
8765966 - 财政年份:2014
- 资助金额:
$ 22.13万 - 项目类别:
Chemical Inhibitors to Define an Essential M. Tuberculosis Signaling Network
化学抑制剂定义重要的结核分枝杆菌信号网络
- 批准号:
8281798 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
Chemical Inhibitors to Define an Essential M. Tuberculosis Signaling Network
化学抑制剂定义重要的结核分枝杆菌信号网络
- 批准号:
8666913 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
Stress-potentiated screening to identify novel inhibitors of M. tuberculosis
应激强化筛选以确定结核分枝杆菌的新型抑制剂
- 批准号:
8500180 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
Stress-potentiated screening to identify novel inhibitors of M. tuberculosis
应激强化筛选以确定结核分枝杆菌的新型抑制剂
- 批准号:
8383145 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
Chemical Inhibitors to Define an Essential M. Tuberculosis Signaling Network
化学抑制剂定义重要的结核分枝杆菌信号网络
- 批准号:
8460462 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
Chemical Inhibitors to Define an Essential M. Tuberculosis Signaling Network
化学抑制剂定义重要的结核分枝杆菌信号网络
- 批准号:
8637915 - 财政年份:2012
- 资助金额:
$ 22.13万 - 项目类别:
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