Development of a Versatile Protein Labeling Tool for Live-Cell Imaging Using Fluorescent β-Lactamase Inhibitors |
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Authors: | Dr. Masafumi Minoshima Taro Umeno Kohei Kadooka Margaux Roux Namiko Yamada Prof. Dr. Kazuya Kikuchi |
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Affiliation: | 1. Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1, Yamadaoka, Suita, Osaka, 5650871 Japan;2. Department of Chemistry, École normale supérieure de Lyon, 15 parvis René Descartes, 69342 Lyon, France |
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Abstract: | To understand the function of protein in live cells, real-time monitoring of protein dynamics and sensing of their surrounding environment are important methods. Fluorescent labeling tools are thus needed that possess fast labeling kinetics, high efficiency, and long-term stability. We developed a versatile chemical protein-labeling tool based on fluorophore-conjugated diazabicyclooctane β-lactamase inhibitors (BLIs) and wild-type TEM-1 β-lactamase protein tag. The fluorescent probes efficiently formed a stable carbamoylated complex with β-lactamase, and the labeled proteins were visualized over a long period of time in live cells. Moreover, use of an α-fluorinated carboxylate ester-based BLI prodrug enabled the probe to permeate cell membranes and stably label intracellular proteins after unexpected spontaneous ester hydrolysis. Lastly, combining the labeling tool with a pH-activatable fluorescent probe allowed visual monitoring of lysosomal protein translocation during autophagy. |
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Keywords: | Autophagy Fluorescence Imaging Probe Lysosomal Trafficking Protein Labeling β-Lactamase Inhibitor |
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