Abstract: New Engineered Luciferase Reporter from a Deep Sea Shrimp

Researchers get great utility from bioluminescent tools due to greater sensitivity, broad dynamic range and higher signal-to-noise ratios. Incremental enhancements of native enzymes including firefly and Renilla luciferase led to many of the improvements. Researchers have sought smaller and brighter luciferases to allow continued development of new applications. In this paper published in ACS Chemical Biology, the authors describe the engineering of both an enzyme and substrate combined to create a novel highly efficient bioluminescence system.

NanoLuc™ Luciferase, a small luciferase subunit (19kDa) from the deep sea shrimp Oplophorus gracilirostris, allows ∼2.5 millionfold improved luminescence activity in mammalian cells by merging optimization of protein structure with development of a novel imidazopyrazinone substrate (furimazine). The glow-type luminescence (signal half-life >2 hours) produced by the new luciferase has a specific activity ∼150-fold greater than either firefly (Photinus pyralis) or Renilla luciferases similarly configured for glow-type assays. In mammalian cells, NanoLuc™ Luciferase shows no evidence of posttranslational modifications or subcellular partitioning. The enzyme exhibits high physical stability, retaining activity with incubation up to 55°C or in culture medium for >15 hours at 37°C.

The authors discuss utility of NanoLuc™ Luciferase as a genetic reporter configured for high sensitivity or for response dynamics through addition of a degradation sequence to reduce intracellular accumulation. Data shows the effect of adding a signal sequence to allow export of NanoLuc™ Luciferase to the culture medium, allowing measurement of enzyme activity without cell lysis.