We have recently developed a genetically encoded fluorescent sensor, named OHSor, that is highly specific to Organic hydroperoxides (OHP) and OHP-generated alkoxy radicals.  In this engineered protein construct, the N and C termini of circularly-permutated fluorescent protein-Venus (cp-Venus) were site-specifically inserted into the regulatory region of an OHP-sensory protein OhrR, which successfully converted the OHP-specific protein conformational changes to an enhanced fluorescence signal.   To demonstrate its applications, we applied OHSor for selectively monitoring of intracellular formation of alkoxy radical in E. coli cells, which led to the discovery that bactericidal antibiotics can drive the intracellular production of highly toxic alkoxyl radicals that may contribute to antibiotic-induced cell death.  The utilization of OHSor for intracellular imaging of lipid oxidation will also be demonstrated in mammalian cells.  Taken together, OHSor may serve as a valuable tool dissecting the physiological and/or pathological roles of alkoxyl radicals in diverse prokaryotic and eukaryotic cells.

Protein chemistry and engineering for live-cell imaging

243rd National Meeting (2012)

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