No-Wash Protein Labeling with Designed Fluorogenic Probes and Application to Real-Time Pulse-Chase Analysis

被引：72

作者：

Mizukami, Shin ^{[1
,2
]}

Watanabe, Shuji ^{[1
]}

Akimoto, Yuri ^{[1
]}

Kikuchi, Kazuya ^{[1
,2
]}

机构：

[1] Osaka Univ, Grad Sch Engn, Div Adv Sci & Biotechnol, Suita, Osaka 5650871, Japan

[2] Osaka Univ, Immunol Frontier Res Ctr IFReC, Suita, Osaka 5650871, Japan

来源：

JOURNAL OF THE AMERICAN CHEMICAL SOCIETY | 2012年 / 134卷 / 03期

基金：

日本科学技术振兴机构; 日本学术振兴会;

关键词：

LACTAMASE-TAG TECHNOLOGY; BETA-LACTAMASE; IN-VIVO; SMALL MOLECULES; LIVING CELLS; CEPHALOSPORIN; REAGENT;

D O I：

10.1021/ja208290f

中图分类号：

O6 [化学];

学科分类号：

0703 ;

摘要：

Small molecule labeling techniques for cellular proteins under physiological conditions are very promising for revealing new biological functions. We developed a no-wash fluorogenic labeling system by exploiting fluorescence resonance energy transfer (FRET)-based fluorescein-cephalosporin-azopyridinium probes and a mutant beta-lactamase tag. Fast quencher elimination, hydrophilicity, and high resistance against autodegradation were achieved by rational refinement of the structure. By applying the probe to real-time pulse-chase analysis, the trafficking of epidermal growth factor receptors between cell surface and intracellular region was imaged. In addition, membrane-permeable derivatization of the probe enabled no-wash fluorogenic labeling of intracellular proteins.

引用

页码：1623 / 1629

页数：7

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