HaloTag-Based Hybrid Targetable and Ratiometric Sensors for Intracellular Zinc
| Autor: | Melissa L. Zastrow, Zhen Huang, Stephen J. Lippard |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
chemistry.chemical_element Intracellular zinc Zinc Endoplasmic Reticulum Nitric Oxide Transfection 01 natural sciences Biochemistry 03 medical and health sciences Humans Nitric Oxide Donors Fluorescent Dyes Cell Nucleus 010405 organic chemistry Chemistry General Medicine Fluoresceins Combinatorial chemistry 0104 chemical sciences Mitochondria Luminescent Proteins 030104 developmental biology Hydrazines Microscopy Fluorescence Molecular Medicine HeLa Cells |
| Zdroj: | ACS chemical biology. 15(2) |
| ISSN: | 1554-8937 |
| Popis: | We report a new series of small molecule-protein hybrid zinc sensors that combine genetic targetability with the spectroscopic profile of synthetic fluorophores. We functionalized the zinc sensor ZinPyr-1 (ZP1) with a chloroalkane linker (ZP1-12Cl) that reacts specifically with the engineered protein HaloTag. The resulting construct, ZP1-HaloTag, binds zinc ions with a threefold fluorescence enhancement. Through exploitation of the protein synthesis machinery of live cells, the HaloTag protein component was expressed, and the ZP1-HaloTag hybrid was assembled upon bath application of ZP1-12Cl. After fusion of HaloTag with targeting peptides or proteins, the resulting hybrid sensor could be directed to specific subcellular locales, including the nucleus, mitochondrial outer membrane, and endoplasmic reticulum. Furthermore, HaloTag was linked with the red fluorescent protein mCherry, permitting formation of a two-fluorophore system that provides not only targetable but also ratiometric sensing of cellular zinc. This system reversibly detects both exogenous and endogenous mobile Zn |
| Databáze: | OpenAIRE |
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