Creation of artificial luciferases for bioassays.

The present study demonstrates the creation of artificial luciferases (ALuc) for bioassays, inspired by a sequence alignment of copepod luciferases. Extraction of the consensus amino acids from the alignment enabled us to generate a series of ALucs with unique optical properties and sequential identities that are clearly different from those of any existing copepod luciferase. For example, some ALucs exhibited heat stability, dramatically prolonged optical intensities, broad full width at half-maximum, and strong optical intensities. The practical suitability of the luciferases as an optical readout was examined in diverse bioassays, including mammalian two-hybrid assays, live cell imaging, single-chain probes, bioluminescent capsules, and bioluminescent antibodies. We further determine the physical properties of ALucs through bioinformatic analysis and finally discuss detailed issues on the unique properties of ALucs. The present study shows how to create the artificial enzymes with excellent optical properties for bioassays and encourages researchers to fabricate their own unique artificial enzymes with designed properties and functionalities.