Four new bright members of the ZinPyr zinc fluorescence sensor family for live cell imaging

As second most abundant trace element, zinc plays numerous roles in the human body. Not only as structural component and co-factor of more than 3,000 proteins, but also as so-called mobile Zn, mZn – pools of labile bound zinc ions. This mZn occurs especially in the central nervous system, where it plays a fundamental role in signal transduction. Accordingly, a dysregulated zinc homeostasis is linked to the pathogenesis of neurodegenerative diseases. Fluorescence sensors have emerged as powerful tool to unravel its role on the molecular level. With 20 members, the most prominent sensor family is the ZinPyr family that exploits a fluorescein platform equipped with bis(2-pyridylmethyl)amine (DPA) as zinc binding unit. Within this article, we report four new bright members of the ZinPyr family, ZP1(5-en), ZP1(6-en), ZP1(5-Me2en), and ZP1(6-Me2en), which are derived from the known sensors ZP1(5-CO2H) and ZP1(6-CO2H). Modification of these parent sensors with ethane-1,2-diamine (en) or N1,N2-dimethylethane-1,2-diamine (Me2en) yielded cell-permeate sensors that combine the low quantum yield of the zinc-free state Φfree (0.165(0) – 0.190(9)) of the parent sensors with a high turn-on (5) and dynamic range (4.2 – 5.4). These properties make the new sensors among the brightest sensors in the ZinPyr family. Live cell imaging proves their ability to detect intracellular zinc with an approximate turn-on of 2-3. The sensors localize presumably in the lysosomes, with ZP1(6-en) and ZP1(5-Me2en) also localizing in the nuclei.