视网膜关利酸环酶激活蛋白5 (GCAP5) 的NMR结构,具有R22A突变,可消除二分化并增强环酶激活
Diana L Cudia1, Effibe O Ahoulou1, Aritra Bej1
1Department of Chemistry, University of California, Davis, Davis, California 95616, United States.
Biochemistry
|April 25, 2024
概括
斑马鱼光受体中的酸环酸激活蛋白-5 (GCAP5) 的突变破坏了二分化,导致视网膜酸环酸 (GC-E) 的增强激活. 这种结构变化改变了结合,增加了GC-E亲和力,解释了活性增加.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 结构生物学 结构生物学
背景情况:
- 关酸环酶激活蛋白-5 (GCAP5) 对于激活斑马鱼光受体中的膜受体视网膜关酸环酶 (GC-E) 至关重要.
- 之前在GCAP5 (GCAP5R22A) 中发现的一种R22A突变被证明可以消除GCAP5的二分化,并与野生型GCAP5 (GCAP5WT) 相比显著增加GC-E激活.
研究的目的:
- 阐明GCAP5中的R22A突变导致二分化亲和力降低和GC-E激活增强的分子机制.
- 研究R22A突变对GCAP5-相互作用和GCAP5-GC-E结合的结构和功能影响.
主要方法:
- 异热定位热量计 (ITC) 用于量化结合亲缘关系.
- 核磁共振 (NMR) 光谱法以确定GCAP5R22A的3D结构.
- 功能循环酶试验测量GC-E激活和结合亲和力.
主要成果:
- ITC揭示了GCAP5R22A与GCAP5WT不同,它结合3个Ca2+离子,其中有两个高亲和度的纳米分子位点可能受到R22A突变的影响.
- 核磁共振结构显示GCAP5R22A与GCAP5WT相似,但与R22A附近的局部结构变化以及C端螺旋方向的改变.
- 在GCAP5R22A中缺乏GCAP5WT中存在的盐桥,这有助于减弱二元化. 结构建模表明R22A增强了与GC-E的疏水相互作用,使结合亲和力增加了约25%.
结论:
- 在GCAP5中R22A突变通过消除关键的盐桥来破坏二分化,并改变结合,导致构成性活性状态.
- GCAP5R22A和GC-E之间增强的疏水性相互作用导致结合亲和度的增加,并随后增强了循环酶活性.
- 这些发现为GCAP5的结构功能关系及其对视觉信号通路的调节提供了关键的见解.
相关概念视频
Activation and Inactivation of G Proteins
7.1K
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...
7.1K
GPCRs Regulate Adenylyl Cylase Activity
5.5K
Some GPCRs transmit signals through adenylyl cyclase (AC), a transmembrane enzyme. AC helps synthesize second messenger cyclic adenosine monophosphate (cAMP). AC catalyzes cyclization reaction and converts ATP to cAMP by releasing a pyrophosphate. The pyrophosphate is further hydrolyzed to phosphate by the enzyme pyrophosphatase, which drives cAMP synthesis to completion. However, cAMP is rapidly degraded to 5′ AMP by the enzymes phosphodiesterase (PDE), preventing overstimulation of...
5.5K
G Protein-coupled Receptors
11.9K
G Protein-Coupled Receptors or GPCRs are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to sensory stimuli such as light, odors, hormones, cytokines, or neurotransmitters.
GPCRs are also called heptahelical, 7TM, or serpentine receptors, and consist of seven (H1-H7) transmembrane alpha-helices that span the bilayer to form a cylindrical core. The transmembrane helices are connected by three extracellular loops and three...
GPCRs are also called heptahelical, 7TM, or serpentine receptors, and consist of seven (H1-H7) transmembrane alpha-helices that span the bilayer to form a cylindrical core. The transmembrane helices are connected by three extracellular loops and three...
11.9K
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
Transducer Mechanism: G Protein–Coupled Receptors
1.9K
G Protein–Coupled Receptors (GPCRs) are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to various stimuli. GPCRs regulate critical physiological pathways and are excellent drug targets for treating diseases such as diabetes, cancer, obesity, depression, or Alzheimer's. Nearly 35% of approved drugs implement their therapeutic effects by selectively interacting with specific GPCRs.
GPCRs are also called heptahelical,...
GPCRs are also called heptahelical,...
1.9K


