在Dictyostelium中评估Ras/Rap GTPases的活性
Stephen F Smith1, Pascale G Charest2,3
1Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ, USA.
Methods in molecular biology (Clifton, N.J.)
|July 2, 2024
概括
研究人员开发了新的生化和活细胞成像方法来测量Ras和Rap小GTPase在Dictyostelium中的活性. 这些测试推进了对这些关键分子开关在真核细胞生物学中的研究.
科学领域:
- 细胞生物学 细胞生物学
- 分子生物学分子生物学
- 生物化学 生化学
背景情况:
- 拉斯和拉普小GTPases是细胞信号通路的关键调节者.
- 研究Dictyostelium中的这些蛋白质为我们提供了关于真核生物学的见解.
- 现有的Ras/Rap活性测定方法很有用,但可以改进.
研究的目的:
- 描述用于评估Dictyostelium中的Ras/Rap活性的新方法.
- 为研究Ras/Rap蛋白的调节和细胞作用提供工具.
- 为了使生物化学和活细胞成像分析GTPase活动.
主要方法:
- 使用Ras结合域的生物化学拉下测试.
- 使用光记者对活细胞进行成像,以监测GTPase活动.
- 这些方法应用于Dictyostelium研究.
主要成果:
- 建立了可靠的方法来量化Ras/Rap GTPase活动.
- 证明了拉下测试对生物化学分析的有用性.
- 验证的光报告器用于实时细胞成像GTPase功能.
结论:
- 描述的方法为研究Ras/Rap信号提供了强大的方法.
- 这些技术有助于更深入地了解GTPase调节和真核细胞中的功能.
- 这项研究为Dictyostelium研究社区提供了有价值的工具.
更多相关视频
13:51Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay
Published on: November 11, 2018
9.8K
09:40Imaging G-protein Coupled Receptor GPCR-mediated Signaling Events that Control Chemotaxis of Dictyostelium Discoideum
Published on: September 20, 2011
18.0K
相关概念视频
Small GTPases - Ras and Rho
3.9K
Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
Three regulatory proteins control their activity:
3.9K
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
Activation and Inactivation of G Proteins
7.0K
Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high...
7.0K
The Ras Gene
6.2K
The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a...
Ras is a...
6.2K
GTPases and their Regulation
8.3K
Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒ small G-proteins consisting of a single domain and large multi-domain G-proteins.
Large G-proteins,...
Large G-proteins,...
8.3K
Rab Cascades
2.6K
Rab GTPases act in a regulated cascade during membrane fusion, helping the lipid bilayers mix. The Rab family of proteins are active when bound to GTP, and inactive when bound to GDP. Hence, they act as guanine nucleotide-dependent molecular switches. Rab-GTP recognizes and binds to long or short-range tethering proteins to capture the target vesicle. These tethers coordinate with SNAREs on the vesicle and the target membrane to assemble the trans SNARE complex that locks the mixing bilayers.
2.6K
