邻近标记(proximity labeling, PL)技术的基本原理是将一个具有连接酶活性的酶(目前植物中广泛应用的为TurboID)与目的蛋白融合, 在连接酶的催化作用下将小分子底物(如生物素(biotin))连接到与目的蛋白邻近的内源蛋白或RNA, 而后可通过富集被标记的蛋白或RNA分别进行质谱分析或RNA-Seq, 可鉴定目的蛋白邻近的蛋白或RNA...
Proximity Labeling(PL),邻近标记 原理 将一个具有邻近标记功能的酶(PL ligase)与目标蛋白(诱饵蛋白,Bait protein)融合,通过酶催化的共价修饰将邻近的蛋白标记上生物素(biotin),最后通过亲和素磁珠富集生物素标记蛋白进行质谱鉴定,分析目的蛋白的互作或邻近的蛋白信息。 Mathew B, et al. Mol Cell Proteomics. (202...
临近标记法互作蛋白检测(Proximity labeling,PL),是一种通过融合外源的酶基因到诱饵蛋白基因,然后催化生成可扩散的小分子,来标记细胞内源性的临近蛋白质,然后通过富集、高分辨率质谱或核酸测序来检测鉴定与靶蛋白互作的蛋白质与核酸。相比传统免疫共沉淀(co-IP)与pull down法,临近标记法检测互作蛋白的最大优势就是能...
Proximity labeling (PL) 临近标记检测临近标记法互作蛋白检测(Proximity labeling,PL) 临近标记法互作蛋白检测(Proximity labeling,PL),是一种通过融合外源的酶基因到诱饵蛋白基因,然后催化生成可扩散的小分子,来标记细胞内源性的临...查看详细>>关于怡科
In comparison to the conventional methods to probe the PPIs, the recently described proximity labeling (PL) approach combined with mass spectrometry (MS)-based quantitative proteomics has emerged as a powerful approach to characterize PPIs. However, the application of PL in planta remains at infancy...
TurboID邻近标记(Proximity labeling, PL)的原理是将TurboID生物素连接酶与诱饵蛋白融合,当诱饵蛋白与靶标蛋白发生互作时,TurboID生物素连接酶通过催化的共价修饰将邻近的蛋白标记上生物素,最后通过亲和素磁珠富集生物素标记的蛋白进行质谱鉴定,从而获得靶标蛋白信息。
Metrics Abstract Many biological processes are executed and regulated through the molecular interactions of proteins and nucleic acids. Proximity labeling (PL) is a technology for tagging the endogenous interaction partners of specific protein ‘baits’, via genetic fusion to promiscuous enzymes that catal...
current PL methods require over 18 h of labeling time or utilize chemicals with limited cell permeability or high toxicity. We used yeast display-based directed evolution to engineer two promiscuous mutants of biotin ligase, TurboID and miniTurbo, which catalyze PL with much greater efficiency than...
Enzyme-catalyzed proximity labeling (PL) has emerged as a new approach to study the spatial and interaction characteristics of proteins in living cells. However, current PL methods require over 18 h of labeling time or utilize chemicals with limited cell permeability or high toxicity. We used ...
Therefore, thorough functional and localization assays are imperative to ensure the fidelity and function of PL enzyme fusion constructs. Consequently, achieving optimal spatial resolution and specificity, while mitigating background labeling, requires meticulous experimental design and validation.172,173 Proxi...