相关实验视频

Updated: Jun 30, 2025

Characterization of Neuronal Lysosome Interactome with Proximity Labeling Proteomics

Characterization of Neuronal Lysosome Interactome with Proximity Labeling Proteomics

Published on: June 23, 2022

探索接近依赖生物化实验的选择:标记酶和亲和净化树脂的比较分析

Karl J Schreiber¹, Eileigh Kadijk¹, Ji-Young Youn^1,2

¹Program in Molecular Medicine, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.

Journal of proteome research

|March 20, 2024

概括

靠近性依赖生物化 (PDB) 实验比较miniTurbo和APEX2酶以及不同类型的链状维丁树脂,揭示了优化蛋白相互作用研究的关键变量. 结果指导改进了PDB实验设计.

科学领域:

分子生物学分子生物学
蛋白质组学是指蛋白质组学.
生物化学生物化学

背景情况:

近距离依赖生物化 (PDB) 技术绘制了蛋白质邻居的地图,有助于功能和定位研究.
优化PDB需要了解标签酶和净化树脂对结果的影响.

研究的目的:

在PDB实验中比较miniTurbo和APEX2标记酶的疗效.
为了评估常规的斯特雷普塔维丁赛法与磁性斯特雷普塔维丁树脂用于生物化蛋白质净化的性能.
确定影响PDB数据质量和交互者识别的关键实验变量.

主要方法:

使用miniTurbo和APEX2标记酶对交换反应DNA结合蛋白43kDa (TDP-43) 相互作用体进行比较分析.
评估用于生物化蛋白净化中的斯特雷普塔维丁和磁性斯特雷普塔维丁树脂.
液体染色学-质谱学 (LC-MS/MS) 和SAINTexpress统计分析用于高可靠性互动体识别.

主要成果:

APEX2发现的TDP-43反应堆比miniTurbo更多,但miniTurbo与已知反应堆的重叠性更大.
在小规模的净化中,色树脂比磁性树脂产生了更多的相互作用体.
树脂特异性结合和酸盐与树脂的比率影响蛋白质丰度分布和统计信心.

结论:

标签酶选择和树脂类型都对PDB结果产生重大影响.
实验变量,如树脂选择和结合比,对于优化PDB研究至关重要.
这些发现为靠近依赖生物化实验的经验优化提供了指导.

关键词:

在APEX2中,APEX2是APEX2的第二个版本.生物ID BioID 是一个生物ID.马格雷辛 (MagReSyn) 公司 MS 这就是SAINTexpress的意义.miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo miniTurbo mini 邻近的相互作用.接近性标签的标签.靠近依赖的生物化.

更多相关视频

Split-BioID — Proteomic Analysis of Context-specific Protein Complexes in Their Native Cellular Environment

Split-BioID — Proteomic Analysis of Context-specific Protein Complexes in Their Native Cellular Environment

Published on: April 20, 2018

In Vitro Biochemical Assays using Biotin Labels to Study Protein-Nucleic Acid Interactions

In Vitro Biochemical Assays using Biotin Labels to Study Protein-Nucleic Acid Interactions

Published on: July 17, 2019

相关概念视频

Labeling DNA Probes

Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...

Tagging and Fusion Proteins

Tagging and Fusion Proteins

Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...