Long-Term Monitoring of Enzymatic Activity at the Single-Organelle Level via Reversed Plasmonic Resonance Energy Transfer

Observing and comprehending enzymatic activity in interorganelle communication can yield critical insights into the intricate mechanisms that govern cellular metabolism. However, real-time and long-term monitoring of enzymatic activity at the single-organelle level is challenging, especially during interorganelle communications. Here, we present an optical technique to track oxygen-dependent enzymatic activity in real-time over several hours, concentrating on endosomal–lysosomal interactions at the single-organelle level through reversed plasmonic resonance energy transfer (rPRET). We created urchin-shaped gold nanoparticles as optical nanoantennas by interfacing with resonating black hole quencher molecules to detect azoreductase in individual lysosomes, revealing differences in enzyme activity within cancer and noncancerous cells. Upon entering the lysosome, the rPRET nanoantenna acts as a hypoxic nanoprobe, effectively detecting oxygen signals within the cell. Noninvasive optical monitoring of oxygen-related enzyme activity during prolonged endosome–lysosome interactions enables the identification of differences in enzymatic activities between intact and apoptotic cells. The rPRET nanoantenna offers a method for examining enzymatic regulation during interorganelle interactions in cellular metabolism, focusing on both single-cell and single-organelle levels.