Sato Shinobu, Ohzawa Shinsuke, Sota Kojiro, Sakamoto Naoto, Udo Ayano, Sueda Shinji, Matsuda Tomoki, Nagai Takeharu, Takenaka Shigeori
Department of Applied Chemistry, Kyushu Institute of Technology, Kitakyushu, Japan.
Department of Bioscience and Bioinformatics, Kyushu Institute of Technology, Iizuka, Japan.
Front Chem. 2022 Jul 8;10:922094. doi: 10.3389/fchem.2022.922094. eCollection 2022.
Potassium-sensing oligonucleotide, PSO, a conjugate of a quadruplex structure-forming oligonucleotide with a peptide incorporating a Förster Resonance Energy Transfer (FRET) chromophore pair, has been developed for fluorescent detection of potassium ion (K) in aqueous medium. PSO could be introduced into cells for real-time imaging of cytoplasmic K concentrations. To perform fluorescent imaging of K on the cell surface, we synthesized twelve PSO derivatives with different types of peptide types and lengths, and oligonucleotide sequences including thrombin-binding aptamer (TBA) sequences with FAM and TAMRA as a FRET chromophore pair, and evaluated their performance. was shown to respond selectively to K, not to most ions present , and to show reciprocal fluorescence changes in response to K concentration. For the peptide chains and oligonucleotide sequences examined in this study, the PSO derivatives had values for K in the range of 5-30 mM. All PSO derivatives showed high K selectivity even in the presence of excess Na. The PSO derivatives were successfully localized to the cell surface by biotinylated concanavalin A (ConA) or sulfo-NHS-biotin streptavidin (StAv). Fluorescence imaging of extracellular K upon addition of apoptosis inducers was successfully achieved by localized to the cell surface.
钾离子感应寡核苷酸（PSO）是一种由形成四链体结构的寡核苷酸与包含福斯特共振能量转移（FRET）发色团对的肽共轭而成的物质，已被开发用于在水性介质中对钾离子（K⁺）进行荧光检测。PSO可被引入细胞中以实时成像细胞质中的K⁺浓度。为了在细胞表面对K⁺进行荧光成像，我们合成了十二种具有不同肽类型和长度以及寡核苷酸序列的PSO衍生物，这些序列包括带有FAM和TAMRA作为FRET发色团对的凝血酶结合适体（TBA）序列，并评估了它们的性能。结果表明，PSO对K⁺具有选择性响应，而对大多数存在的离子无响应，并且会随着K⁺浓度的变化呈现出相反的荧光变化。对于本研究中检测的肽链和寡核苷酸序列，PSO衍生物对K⁺的解离常数（Kd）值在5 - 30 mM范围内。即使在存在过量Na⁺的情况下，所有PSO衍生物仍表现出高K⁺选择性。通过生物素化的伴刀豆球蛋白A（ConA）或磺基-NHS-生物素 - 链霉亲和素（StAv），PSO衍生物成功定位于细胞表面。通过定位于细胞表面，成功实现了在添加凋亡诱导剂后对细胞外K⁺的荧光成像。
