Protein-specific localization of a rhodamine-based calcium-sensor in living cells

Marcel Best; Isabel Porth; Sebastian Hauke; Felix Braun; Dirk-Peter Herten; Richard Wombacher

doi:10.1039/C6OB00365F

Protein-specific localization of a rhodamine-based calcium-sensor in living cells

Marcel Best, Isabel Porth, Sebastian Hauke, Felix Braun, Dirk-Peter Herten, Richard Wombacher

Cardiovascular Sciences

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Abstract

A small synthetic calcium sensor that can be site-specifically coupled to proteins in living cells by utilizing the bio-orthogonal HaloTag labeling strategy is presented. We synthesized an iodo-derivatized BAPTA chelator with a tetramethyl rhodamine fluorophore that allows further modification by Sonogashira cross-coupling. The presented calcium sensitive dye shows a 200-fold increase in fluorescence upon calcium binding. The derivatization with an aliphatic linker bearing a terminal haloalkane-function by Sonogashira cross-coupling allows the localization of the calcium sensor to Halo fusion proteins which we successfully demonstrate in in vitro and in vivo experiments. The herein reported highly sensitive tetramethyl rhodamine based calcium indicator, which can be selectively localized to proteins, is a powerful tool to determine changes in calcium levels inside living cells with spatiotemporal resolution.

Original language	English
Pages (from-to)	5606-5611
Journal	Organic and Biomolecular Chemistry
Volume	14
DOIs	https://doi.org/10.1039/C6OB00365F
Publication status	Published - 7 Apr 2016

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title = "Protein-specific localization of a rhodamine-based calcium-sensor in living cells",

abstract = "A small synthetic calcium sensor that can be site-specifically coupled to proteins in living cells by utilizing the bio-orthogonal HaloTag labeling strategy is presented. We synthesized an iodo-derivatized BAPTA chelator with a tetramethyl rhodamine fluorophore that allows further modification by Sonogashira cross-coupling. The presented calcium sensitive dye shows a 200-fold increase in fluorescence upon calcium binding. The derivatization with an aliphatic linker bearing a terminal haloalkane-function by Sonogashira cross-coupling allows the localization of the calcium sensor to Halo fusion proteins which we successfully demonstrate in in vitro and in vivo experiments. The herein reported highly sensitive tetramethyl rhodamine based calcium indicator, which can be selectively localized to proteins, is a powerful tool to determine changes in calcium levels inside living cells with spatiotemporal resolution.",

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T1 - Protein-specific localization of a rhodamine-based calcium-sensor in living cells

AU - Best, Marcel

AU - Porth, Isabel

AU - Hauke, Sebastian

AU - Braun, Felix

AU - Herten, Dirk-Peter

AU - Wombacher, Richard

PY - 2016/4/7

Y1 - 2016/4/7

N2 - A small synthetic calcium sensor that can be site-specifically coupled to proteins in living cells by utilizing the bio-orthogonal HaloTag labeling strategy is presented. We synthesized an iodo-derivatized BAPTA chelator with a tetramethyl rhodamine fluorophore that allows further modification by Sonogashira cross-coupling. The presented calcium sensitive dye shows a 200-fold increase in fluorescence upon calcium binding. The derivatization with an aliphatic linker bearing a terminal haloalkane-function by Sonogashira cross-coupling allows the localization of the calcium sensor to Halo fusion proteins which we successfully demonstrate in in vitro and in vivo experiments. The herein reported highly sensitive tetramethyl rhodamine based calcium indicator, which can be selectively localized to proteins, is a powerful tool to determine changes in calcium levels inside living cells with spatiotemporal resolution.

AB - A small synthetic calcium sensor that can be site-specifically coupled to proteins in living cells by utilizing the bio-orthogonal HaloTag labeling strategy is presented. We synthesized an iodo-derivatized BAPTA chelator with a tetramethyl rhodamine fluorophore that allows further modification by Sonogashira cross-coupling. The presented calcium sensitive dye shows a 200-fold increase in fluorescence upon calcium binding. The derivatization with an aliphatic linker bearing a terminal haloalkane-function by Sonogashira cross-coupling allows the localization of the calcium sensor to Halo fusion proteins which we successfully demonstrate in in vitro and in vivo experiments. The herein reported highly sensitive tetramethyl rhodamine based calcium indicator, which can be selectively localized to proteins, is a powerful tool to determine changes in calcium levels inside living cells with spatiotemporal resolution.

U2 - 10.1039/C6OB00365F

DO - 10.1039/C6OB00365F

M3 - Article

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EP - 5611

JO - Organic and Biomolecular Chemistry

JF - Organic and Biomolecular Chemistry

ER -

Protein-specific localization of a rhodamine-based calcium-sensor in living cells

Abstract

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