相关实验视频
Updated: Jul 6, 2026

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Measuring In Vitro ATPase Activity for Enzymatic Characterization
Published on: August 23, 2016
一个具有强烈和特异性的ATP三酸盐识别能力的小体
Peter L Sazani1, Rosa Larralde, Jack W Szostak
1Howard Hughes Medical Institute and Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
Journal of the American Chemical Society
|July 9, 2004
概括
研究人员开发了一种新型RNA吸收酶,通过识别其5'酸盐,专门与腺三酸盐 (ATP) 结合. 这种胺体将ATP与AMP区分开来,具有1000倍以上的选择性.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 在RNA Aptamer技术上
背景情况:
- RNA 亚体是被选中的核酸分子,专门用于与标结合.
- 之前的ATP体选择往往缺乏对联体的酸化状态的特异性.
- 区分腺酸盐对于理解细胞能量代谢至关重要.
研究的目的:
- 开发一种能够根据它们的5'酸化状态来区分腺素连接体的RNA吸附体.
- 创建一个aptamer,专门识别ATP的三酸盐组.
- 研究小RNA与充电部分相互作用的潜力.
主要方法:
- 实验室内选择 (SELEX) 用于分离RNA吸附体.
- 施加了选择性压力,以支持能识别ATP的5'三酸盐的aptamer.
- 用生物物理方法确定了ATP和AMP的结合亲和度 (Kd).
主要成果:
- 成功选择了一种具有高ATP特异性的RNA胺体.
- 胺体表现出ATP (Kd ≈ 5 μM) 和AMP (Kd ≈ 5.5 mM) 之间的结合亲和力差异为1100倍.
- 这表明,阿普坦体能够根据5'酸化状态进行区分.
结论:
- 小RNA分子可以被设计成与酸盐等负电荷部分相互作用并识别它们.
- 选择的胺体为研究与ATP相关的生物过程提供了有价值的工具.
- 这项工作突显了RNA吸收体在分子识别应用中的多功能性.
相关概念视频
ATP Driven Pumps I: An Overview
ATP-driven pumps, also known as transport ATPases, are integral membrane proteins. They have binding sites for ATP located on the membrane's cytosolic side and the ion-conducting domain in the transmembrane region. These pumps use the free energy released from ATP hydrolysis to move the solutes across cell membranes against an electrochemical gradient.
There are four main types of ATP-driven pumps - P-type, V-type, F-type, and ABC transporter. All these pumps are of varying complexities and are...
There are four main types of ATP-driven pumps - P-type, V-type, F-type, and ABC transporter. All these pumps are of varying complexities and are...
ATP Synthase: Structure
ATP synthase or ATPase is among the most conserved proteins found in bacteria, mammals, and plants. This enzyme can catalyze a forward reaction in response to the electrochemical gradient, producing ATP from ADP and inorganic phosphate. ATP synthase can also work in a reverse direction by hydrolyzing ATP and generating an electrochemical gradient. Different forms of ATP synthases have evolved special features to meet the specific demands of the cell. Based on their specific feature, ATP...
The ADP/ATP Carrier Protein
ADP/ATP carrier or AAC protein is the most abundant carrier protein in the inner mitochondrial membrane. It transports large quantities of ADP and ATP, equivalent to the average human body weight, every day. Among other transporters, ACC protein is one of the best-studied members of the mitochondrial carrier protein family. The ADP/ATP carrier protein comprises two transmembrane helices connected to a loop and a single alpha-helix on the matrix side. It switches between two conformational...
ATP Driven Pumps II: P-type Pumps
The P-type pumps are a large family of integral membrane transporter ATPases. They are divided into five major types based on substrate specificity, from I to V.
A typical P-type pump has three cytosolic domains: nucleotide-binding (N), phosphorylation (P), and activator (A) domains. These domains are connected to the membrane-spanning helices by short amino acid segments. ATP hydrolysis and covalent phosphoenzyme intermediate formation are crucial parts of the catalytic cycle. At the highly...
A typical P-type pump has three cytosolic domains: nucleotide-binding (N), phosphorylation (P), and activator (A) domains. These domains are connected to the membrane-spanning helices by short amino acid segments. ATP hydrolysis and covalent phosphoenzyme intermediate formation are crucial parts of the catalytic cycle. At the highly...
ATP Driven Pumps III: V-type Pumps
V-type pumps are ATP-driven pumps found in the vacuolar membranes of plants, yeast, endosomal and lysosomal membranes of animal cells, plasma membranes of a few specialized eukaryotic cells, and some prokaryotes. They are also known as the V1Vo-ATPase, that couple ATP hydrolysis to transport protons against a concentration gradient.
The peripheral or cytosolic V1 domain with eight subunits is involved in ATP hydrolysis. The integral or transmembrane V0 domain containing at least five subunits...
The peripheral or cytosolic V1 domain with eight subunits is involved in ATP hydrolysis. The integral or transmembrane V0 domain containing at least five subunits...
Structure of Porins
Mitochondria, chloroplasts, and gram-negative bacteria have transmembrane, beta-barrel proteins called porins to mediate the free diffusion of ions and metabolites across the membrane. Mitochondrial porin precursors contain conserved amino acid sequences called beta signals at their C-terminal. Beta signals have a motif of PoXGXXHyXHy (Po-Polar, X-Any amino acid, G-Glycine, Hy-LargeHydrophobic), which are crucial for precursor recognition to initiate precursor assembly. Beta-barrel precursors...

