Notes: Researchers used NanoLuc® Luciferase technology to monitor the interaction between and LDLR and PCSK9. (4931)

Notes: These authors describe the engineering of an enzyme and substrate to create a novel highly efficient bioluminescence system. The paper introduces NanoLuc™ Luciferase, a small luciferase subunit (19kDa) from the deep-sea shrimp Oplophorus gracilirostris, which provides ∼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 or Renilla luciferases similarly configured for glow-type assays. In mammalian cells, NanoLuc™ Luciferase shows no evidence of post-translational 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. (4295)