Singlet oxygen generation by the genetically encoded tag miniSOG.

Area of Science:

• Biochemistry
• Microscopy
• Photochemistry

Background:

• Genetically encodable fluorescent tags are crucial for advanced microscopy techniques.
• MiniSOG is a fluorescent protein tag known for singlet oxygen production upon light irradiation.
• Previous studies reported a high quantum yield (ΦΔ) for miniSOG's singlet oxygen generation.

Purpose of the Study:

• To accurately determine the quantum yield (ΦΔ) of singlet oxygen production by miniSOG.
• To investigate the photochemistry of miniSOG and identify reasons for discrepancies in reported ΦΔ values.
• To explore factors influencing miniSOG's singlet oxygen generation efficiency.

Main Methods:

• Direct measurement of miniSOG's phosphorescence at 1275 nm.
• Chemical trapping of singlet oxygen using uric acid as a probe.
• Analysis of miniSOG's photooxidation of anthracene dipropionic acid (ADPA).

Main Results:

• A significantly lower ΦΔ value (0.03 ± 0.01) was determined using phosphorescence and uric acid trapping.
• MiniSOG was found to oxidize ADPA via both singlet oxygen-dependent and -independent pathways.
• Cumulative light irradiation increased miniSOG's ΦΔ by approximately 10-fold due to protein transformation.

Conclusions:

• The actual quantum yield of singlet oxygen production by miniSOG is lower than previously reported.
• MiniSOG's singlet oxygen generation mechanism is complex, involving both direct and indirect pathways.
• Photoinduced protein transformation enhances miniSOG's efficiency, potentially explaining its superior performance compared to other fluorescent proteins.