Design and synthesis of a FlAsH-type Mg2+ fluorescent probe for specific protein labeling

Tomohiko Fujii, Yutaka Shindo, Kohji Hotta, Daniel Citterio, Shigeru Nishiyama, Koji Suzuki, Kotaro Oka

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Although the magnesium ion (Mg2+) is one of the most abundant divalent cations in cells and is known to play critical roles in many physiological processes, its mobilization and underlying mechanisms are still unknown. Here, we describe a novel fluorescent Mg2+ probe, "KMG-104-AsH", composed of a highly selective fluorescent Mg 2+ probe, "KMG-104", and a fluorescence-recoverable probe, "FlAsH", bound specifically to a tetracysteine peptide tag (TCtag), which can be genetically incorporated into any protein. This probe was developed for molecular imaging of local changes in intracellular Mg2+ concentration. KMG-104-AsH was synthesized, and its optical properties were investigated in solution. The fluorescence intensity of KMG-104-AsH (at λem/max = 540 nm) increases by more than 10-fold by binding to both the TCtag peptide and Mg2+, and the probe is highly selective for Mg2+ (Kd/Mg = 1.7 mM, Kd/Ca 100 mM). Application of the probe for imaging of Mg2+ in HeLa cells showed that this FlAsH-type Mg2+ sensing probe is membrane-permeable and binds specifically to tagged proteins, such as TCtag-actin and mKeima-TCtag targeted to the cytoplasm and the mitochondrial intermembrane space. KMG-104-AsH bound to TCtag responded to an increase in intracellular Mg2+ concentration caused by the release of Mg2+ from mitochondria induced by FCCP, a protonophore that eliminates the inner membrane potential of mitochondria. This probe is expected to be a strong tool for elucidating the dynamics and mechanisms of intracellular localization of Mg2+.

Original languageEnglish
Pages (from-to)2374-2381
Number of pages8
JournalJournal of the American Chemical Society
Volume136
Issue number6
DOIs
Publication statusPublished - 2014 Feb 12

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Fluorescent Dyes
Labeling
Peptides
Proteins
Mitochondria
Fluorescence
Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone
Physiological Phenomena
Molecular Imaging
Divalent Cations
Membranes
HeLa Cells
Molecular imaging
Membrane Potentials
Magnesium
Actins
Cytoplasm
Ions
1-(2,7-difluoro-6-hydroxy-4,5-bis(1,2,3-dithioarsolan-2-yl)-3-oxo-3H-xanthen-9-yl)-4-oxo-4H-quinolizine-3-carboxylic Acid
Optical properties

ASJC Scopus subject areas

  • Chemistry(all)
  • Catalysis
  • Biochemistry
  • Colloid and Surface Chemistry

Cite this

Design and synthesis of a FlAsH-type Mg2+ fluorescent probe for specific protein labeling. / Fujii, Tomohiko; Shindo, Yutaka; Hotta, Kohji; Citterio, Daniel; Nishiyama, Shigeru; Suzuki, Koji; Oka, Kotaro.

In: Journal of the American Chemical Society, Vol. 136, No. 6, 12.02.2014, p. 2374-2381.

Research output: Contribution to journalArticle

Fujii, Tomohiko ; Shindo, Yutaka ; Hotta, Kohji ; Citterio, Daniel ; Nishiyama, Shigeru ; Suzuki, Koji ; Oka, Kotaro. / Design and synthesis of a FlAsH-type Mg2+ fluorescent probe for specific protein labeling. In: Journal of the American Chemical Society. 2014 ; Vol. 136, No. 6. pp. 2374-2381.
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