Protein conformational changes drive signal transduction to regulate cellular activities, yet monitoring of these changes in living cells remains challenging. Here, we introduce BIOSCE (BIOprobe based on Steric Confinement-induced Emission), a technique that enables tracking of individual protein conformations in living cells across millisecond-to-minute timescales. BIOSCE reports protein conformational changes via steric confinement-induced luminescence switching from non-luminescent to luminescent states. We demonstrate that BIOSCE rapidly senses calmodulin conformational changes triggered by intracellular calcium fluctuations. The BIOSCE platform achieved millisecond-resolution monitoring of single-protein conformations within cellular signaling pathways, as evidenced by its sensitive detection of rapamycin-dependent FKBP (FK506-binding protein)-FRB (FKBP-rapamycin binding) interactions regardless of the labeled partner. Furthermore, we applied BIOSCE to track the spatial distribution of SNAP25 (25 kDa synaptosomal nerve-associated protein) during botulinum neurotoxin A (BoNT/A) intoxication, revealing differential catalytic processing of its cleavage fragments. This generalizable approach provides a robust platform for investigating single-molecule conformational changes with high spatiotemporal resolution and enables direct evaluation of transient cellular events.
Ma et al. (2026) studied this question.