Activity-based Genetically Encoded Fluorescent and Luminescent Probes for Detecting Formaldehyde in Living Cells.

Formaldehyde (FA) is endogenously produced in living systems through a variety of biological processes, and has been implicated in many pathological conditions. Detection tools for biological FA are, therefore, of great interest. Here we report novel activity-based genetically encoded fluorescent and luminescent probes for detecting FA in aqueous solutions and living mammalian cells. A FA-reactive lysine analogue, PrAK, was site-specifically incorporated into the essential lysine sites of enhanced green fluorescent protein (EGFP) and firefly luciferase (fLuc) to afford the fluorescent and luminescent probes, respectively. FA selectively reacts with the PrAK residues on EGFP and fLuc through 2-aza-Cope rearrangement, resulting in fluorescence and luminescence turn-on responses, respectively, to FA over potentially interfering reactive species in aqueous buffer. Moreover, the genetically encoded probes are capable of visualizing FA at physiologically relevant levels in living mammalian cells by fluorescence and luminescence imaging, demonstrating their potential as new tools to explore FA biology.