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...
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
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...
184Citations 107Altmetric Metrics Abstract This protocol describes the use of TurboID and split-TurboID in proximity labeling applications for mapping protein–protein interactions and subcellular proteomes in live mammalian cells. TurboID is an engineered biotin ligase that uses ATP to convert biotin int...
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...
a Identification of proteins that are specifically associated with SUN1 at the INM using proximity labeling. The ONM protein WIT1 serves as a control to exclude proteins associated with both the INM and the ONM. b Pairwise ratiometric analyses of PL-LFQMS data using 35S: HA-BioID2-SUN1 ...
Proximity LabelingMurat Sunbul Andres Jäschke EditorsMethods and ProtocolsMethods in Molecular Biology 2008 阅读了该文档的用户还阅读了这些文档 108 p. ISO 25539-3-2024 18 p. ISO 16810-2024 38 p. ISO 14356-2024 10 p. ISO 15708-1-2024 72 p. BS EN 1982-2024 50 p. PD ISO-...
This protocol describes the use of TurboID and split-TurboID in proximity labeling applications for mapping protein–protein interactions and subcellular proteomes in live mammalian cells. TurboID is an engineered biotin ligase that uses ATP to convert biotin into biotin–AMP, a reactive intermediate th...
APEX is an engineered peroxidase that functions as an electron microscopy tag and a promiscuous labeling enzyme for live-cell proteomics. Because limited sensitivity precludes applications requiring low APEX expression, we used yeast-display evolution to improve its catalytic efficiency. APEX2 is far mor...