Fluorescence energy transfer probes based on the guanine quadruplex formation for the fluorometric detection of potassium ion

Dual-labeled oligonucleotide derivative, FAT-0, carrying 6-carboxyfluorescein (FAM) and 6-carboxy-tetramethylrhodamine (TAMRA) labels at 5′- and 3′-termini of thrombin-binding aptamer (TBA) sequence 5′-GGTTGGTGTGGTTGG-3′ and its derivatives, FAT-n (n = 3, 5, and 7) were designed and synthesized. FAT-n derivatives contained a T_mA spacer (m = 2, 4, and 6, respectively) at 5′-end of TBA sequence. The probes were developed to estimate the spacer effect on FRET efficiency and to identify the best probe for sensing of K⁺. Circular dichroism (CD), UV-vis absorption, and fluorescence studies revealed that all FAT-n probes could form the intramolecular tetraplex structures after binding K⁺. Association constants of particular K⁺/FAT-n complexes were determined using different experimental approaches. Suitability of particular probes for sensitive monitoring of K⁺ in intra- and extracellular conditions was examined and discussed. Calibration graphs of fluorescence ratio were linear in the K⁺ concentration range of 2-10 mM for extracellular conditions showing sensitivity of 1.2% mM⁻¹ K⁺ and for intracellular conditions in the range of 100-200 mM with sensitivity of 0.49% mM⁻¹ K⁺.