A color-tailored fluorogenic sensor for hydrogen peroxide

Abstract

H2O2 is an important signaling molecule and redox regulator of normal cellular metabolism and a major element of oxidative stress. Here we report HyPerFLEX (HyPer with flexible fluorogen excitation), a sensor from the HyPer family designed for high-precision H2O2 monitoring in living cells. HyPerFLEX combines the redox-sensitive OxyR domain from Neisseriameningitidis and circularly permuted fluorogenic protein Y-FAST, yielding oxygen-independent fluorescence upon oxidation of OxyR by H2O2. HyPerFLEX enables imaging H2O2 dynamics in living cells, with tunable spectra from green to far red for multicompartment imaging, even under prolonged hypoxia. It surpasses HyPer7 in detecting ultralow H2O2 concentrations, such as during early glucose-stimulated insulin production, and can measure H2O2 levels in the highly oxidizing endoplasmic reticulum lumen. These advanced features and broad compatibility make HyPerFLEX a powerful tool for studying oxidative stress and cellular signaling. (Figure presented.)