RRC ID 72273
Author Kim SB, Nishihara R, Citterio D, Suzuki K.
Title Fabrication of a New Lineage of Artificial Luciferases from Natural Luciferase Pools.
Journal ACS Comb Sci
Abstract The fabrication of artificial luciferases (ALucs) with unique optical properties has a fundamental impact on bioassays and molecular imaging. In this study, we developed a new lineage of ALucs with unique substrate preferences by extracting consensus amino acids from the alignment of 25 copepod luciferase sequences available in natural luciferase pools. The primary sequence was first created with a sequence logo generator resulting in a total of 11 sibling sequences. Phylogenetic analysis shows that the newly fabricated ALucs form an independent branch, genetically isolated from the natural luciferases, and from a prior series of ALucs produced by our laboratory using a smaller basis set. The new lineage of ALucs were strongly luminescent in living mammalian cells with specific substrate selectivity to native coelenterazine. A single-residue-level comparison of the C-terminal sequences of new ALucs reveals that some amino acids in the C-terminal ends are greatly influential on the optical intensities but limited in the color variance. The success of this approach guides on how to engineer and functionalize marine luciferases for bioluminescence imaging and assays.
Volume 19(9)
Pages 594-599
Published 2017-9-11
DOI 10.1021/acscombsci.7b00081
PMID 28742969
MeSH Amino Acid Sequence Animals COS Cells Cell Line Chlorocebus aethiops Consensus Sequence Copepoda / enzymology* Luciferases / chemistry* Luciferases / genetics Luciferases / metabolism Luminescent Measurements Phylogeny Sequence Alignment Substrate Specificity
DNA material ALuc49 (RDB19617)