在单分子水平上探测囊泡重建β2上腺素受体的激活和形态动力学
Marijonas Tutkus1,2,3, Christian V Lundgaard1, Salome Veshaguri1
1Department of Chemistry, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark.
The journal of physical chemistry. B
|February 23, 2024
概括
单个G蛋白结合受体 (GPCR),如β2上腺素受体 (β2AR),表现出动态的结构变化. 激素结合改变了这些动态,影响了与原生相似的膜环境中的受体信号表型.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 生物物理学的生物物理.
背景情况:
- G-蛋白结合受体 (GPCR) 是关键的膜蛋白,参与许多生理过程.
- 了解GPCR的动态结构行为是解释其功能可塑性和多样化的信号结果的关键.
- 尽管结构生物学取得了进展,但GPCRs的结构格局仍然复杂且不完全理解.
研究的目的:
- 为了监测单个β-2上腺素受体 (β2ARs) 在与原生类似的脂质膜环境中的构造动态.
- 为了研究激素激励如何影响这些单分子动态.
- 用光显微镜来描述β2ARs的构造状态和转变.
主要方法:
- 使用光显微镜测定,特别是总内部反射光显微镜 (TIRFM).
- 采用单个小单性脂质体试验,在蛋白质脂质体中复制β2ARs,而不是洗剂小粒.
- 通过环境敏感的BODIPY光体连接到螺旋的细胞质末端的β2AR来监测构造动力学6.
主要成果:
- 标有BODIPY 493/503的单个β2ARs在第二个时间表上表现出动态强度波动.
- 激素刺激 (BI-167107) 改变了β2AR动态,增加了高强度状态及其持续时间.
- 过渡密度图显示了激动剂诱导的从低/中等到中等/高强度状态的转变,这表明了形状选择.
结论:
- 单分子光显微镜在与原生相似的膜环境中提供了对GPCR结构动态的洞察.
- 对β2ARs的激素结合调节其构造组合,有利于特定的状态.
- 这种方法是在脂质双层内表征GPCR结构动态方面迈出了一个有希望的步骤.
更多相关视频
10:59Visualizing the Conformational Dynamics of Membrane Receptors Using Single-Molecule FRET
Published on: August 17, 2022
3.2K
14:09Fluorescence Biomembrane Force Probe: Concurrent Quantitation of Receptor-ligand Kinetics and Binding-induced Intracellular Signaling on a Single Cell
Published on: August 4, 2015
12.5K
相关概念视频
Pinching-off of Coated Vesicles
3.1K
Vesicle budding is orchestrated by distinct cytosolic proteins such as adaptor proteins, coat proteins, and GTPases. To initiate vesicle budding, membrane-bending proteins containing crescent-shaped BAR domains bind to the lipid heads in the bilayer and distort the membrane to form a protein-coated vesicle bud. Adaptors proteins such as AP2 for clathrin-coated vesicles can nucleate on the deformed membrane. Finally, coat proteins such as clathrin or COPI and COPII assemble into a coat forming...
3.1K
Protein Dynamics in Living Cells
2.1K
Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
2.1K
Rab Proteins
3.9K
Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
3.9K
Clathrin Coated Vesicles
7.0K
Clathrin-coated vesicles use endocytosis to transport receptors and lysosomal hydrolases from the Golgi to the lysosome in the late secretory pathway. Clathrin-mediated endocytosis was the first described endocytic process, and Clathrin-coated vesicles remain one of the most well-studied transport vesicles. The molecular machinery that generates clathrin-coated vesicles comprises over 50 proteins that precisely coordinate vesicle formation. Cell surface receptors concentrated in indented sites...
7.0K
