Part:BBa_K1616023

T7 promoter - HisTag- NanoLuc - T7 terminator

NanoLuc

NanoLuc (Nluc) is a novel engineered luciferase coupled with a novel coelenterazine analogue: Furimazine. When both are coupled, we obtain a much brighter luminescence than Fluc or Rluc and it is known to work exceptionally well as a reporter. NanoLuc is a 19.1 kDa enzyme which is an ATP independent luciferase providing a glow-type luminescence.

Thermal stable enzyme
NanoLuc retains activity following 30 min incubation at 55 °C.
Active over broad pH range
NanoLuc is fully active between pH 7-9 and retains significant activity at pH 5-7
Monomeric enzyme
NanoLuc facilitates use as transcriptional reporter or fusion partner.

A fast detection system involving chemical processes

One other method of fast detection is bioluminescence. This trait found in several marine bacteria emits light by the mean of biochemical mechanisms involving chemical processes. It is important to make the distinction between fluorescence and bioluminescence.In fluorescence, a high energy photon excites a molecule to an excited state which makes the latter lose energy. This loss leads to the release of a visible photon and the drop to the original groundstate of the molecule. Bioluminescence involves an oxygen oxidation of an organic molecule. A luciferase catalyzes the oxidation of luciferin to excited state oxyluciferin which release a visible photon (Shreeram Akilesh, Bioluminescence: Nature’s Bright Idea,2000).

Luciferase: an essential enzyme for bioluminescence

Firefly (Fluc, 61 kDa) and Renilla (Rluc, 36 kDa) luciferases have been used for a huge amount of applications such as for the study of molecular and cellular processes. Given this fact, researchers tried to investigate alternative luciferases from marine organims and achieve a more optimal enzyme structure which could explore other variants of luminogenic substrate. Furthermore, as most of the luciferases come from marine organisms, the substrate the most used is coelenterazine.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal XhoI site found at 117
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 20
  

Design Notes

All BioBrick parts used for assembling the composite part are compatible with the RFC10 Biobrick standard.

Source

NanoLuc is a 19.1 kDa enzyme which is an ATP independent luciferase providing a glow-type luminescence.

References

Kyle Hooper, Ph.D, Applications of a smaller, brighter, more versatile luciferase:NanoLuc™ Luciferase Technology, 2012
Shreeram Akilesh, Bioluminescence: Nature’s Bright Idea,2000.