Live-Cell Imaging of Protein Degradation Utilizing Designed Protein-Tag Mutant and Fluorescent Probe with Turn-Off Switch

Jingchi Gao, Yuichiro Hori, Osamu Takeuchi, Kazuya Kikuchi

Research output: Contribution to journalArticlepeer-review

3 Citations (Scopus)

Abstract

Protein degradation plays various roles in cellular homeostasis and signal transduction. Real-time monitoring of the degradation process not only contributes to the elucidation of relevant biological phenomena but also offers a powerful tool for drug discoveries targeting protein degradation. Fluorescent protein labeling with a protein tag and a synthetic fluorescent probe is a powerful technique that enables the direct visualization of proteins of interest in living cells. Although a variety of protein tags and their labeling probes have been reported, techniques for the visualization of protein degradation in living cells remain limited. In order to overcome this limitation, we herein employed a PYP-tag labeling probe with a fluorescence turn-off switch that enables the imaging of protein degradation. Furthermore, we performed a structure-based design of a PYP-tag to stabilize a complex formed by the probe and the protein tag for long-term live-cell imaging. We successfully applied this technique to live-cell imaging of the degradation process of Regnase-1 in response to immunostimulation.

Original languageEnglish
Pages (from-to)577-583
Number of pages7
JournalBioconjugate Chemistry
Volume31
Issue number3
DOIs
Publication statusPublished - Mar 18 2020
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Pharmacology
  • Pharmaceutical Science
  • Organic Chemistry

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