Cationic Europium Complexes for Visualizing Fluctuations in Mitochondrial ATP Levels in Living Cells

Abstract

The ability to study cellular metabolism and enzymatic processes involving adenosine triphosphate (ATP) is impeded by the lack of imaging probes capable of signalling the concentration and distribution of intracellular ATP rapidly, with high sensitivity. We report here the first example of a luminescent lanthanide complex capable of visualizing changes in the concentration of ATP in the mitochondria of living cells. Four cationic europium(III) complexes [Eu.1–4]+ have been synthesized and their binding capabilities towards nucleoside polyphosphate anions examined in aqueous solution at physiological pH. Complexes [Eu.1]+ and [Eu.3]+ bearing hydrogen bond donor groups in the pendant arms showed excellent discrimination between ATP, ADP and monophosphate species. Complex [Eu.3]+ showed relatively strong binding to ATP (logK a=5.8), providing a rapid, long‐lived luminescent signal that enabled its detection in a highly competitive aqueous medium containing biologically relevant concentrations of Mg2+, ADP, GTP, UTP and human serum albumin. This EuIII complex responds linearly to ATP within the physiological concentration range (1–5 mm), and was used to continuously monitor the apyrase‐catalyzed hydrolysis of ATP to ADP in vitro. We demonstrate that [Eu.3]+ can permeate mammalian (NIH‐3T3) cells efficiently and localize to the mitochondria selectively, permitting real‐time visualization of elevated mitochondrial ATP levels following treatment with a broad spectrum kinase inhibitor, staurosporine, as well as depleted ATP levels upon treatment with potassium cyanide under glucose starvation conditions.

Tuning the anion binding properties of lanthanide receptors to discriminate nucleoside phosphates in a sensing array

“…An exception to this is [Eu.2] + , where the ATP titration in the presence of 5 mM MgCl 2 produced a two-step binding prole, which prevented the determination of a binding constant, due to the occurrence of multiple equilibria involved in the binding of [Eu.2] + to both ATP and ATP-Mg, as indicated by previously reported mass spectral data. 38 Binding 40 This revealed that q ¼ 1 for each complex in the absence of added anions, but q ¼ 0 in the presence of ATP or ADP, consistent with displacement of the coordinated water molecule from each complex upon binding ATP or ADP. In the presence of 5 mM AMP, a q value of 0.3-0.7 was found, indicating partial hydration, possibly reecting the weaker monodentate binding of AMP, in accordance with previous examples of phosphate binding at lanthanide centres.…”

Direct Monitoring of Bicarbonate Transport by Emission Spectroscopy

“…Indeed, a derivative of [Eu.L 1 ] + has already been shown to permeate living cells and localise to specific subcellular compartments. 58 Direct Monitoring of Bicarbonate Transport by Emission Spectroscopy…”

Molecular Receptors for Recognition and Sensing of Nucleotides

“…2019, 25,2684 -2694 www.chemeurj.org ATP, adeninel ocates outsidet he pyrene dimer.P yrene-based Zn II complex 52,p resented by Xing and co-workers, has similar binding and fluorescence properties compared to those found for 51.B ased on the monomer/excimeri ntensity ratio, the selectivity order in a1 0mm HEPES buffer (pH 7.4) of ATP > ADP % GTP > TTP % CTP > AMP > UTP % PPi was observed. [36] Excimer emission is also ak ey feature of the Butler's tripodalr eceptor (60), which can detect ATPa nd quantify apyrase activity. [37] Our group used the anthracene-based DPA-Zn II complex 53 for experimental quantificationo fp-p stackingi nteractions between anthracenea nd nucleobases.…”

“…An improved selectivity for ATPw as achieved with complex 69. [60] This probe can detect changes in concentration of mitochondrial ATP( 1-5 mm)a nd is minimally perturbed by changes in pH, dissolved oxygen, or the presence of proteins. Ta ng and co-workersr eported another turn-onc yclam-based probe (70), which was found to be selective for ATPo ver ADP andA MP.…”