Directed Molecular Evolution Reveals Gaussia Luciferase Variants with Enhanced Light Output Stability

Gaussia Luciferase (Gluc) has proven to be a powerful mammalian cell reporter for monitoring numerous biological processes in immunology, virology, oncology, and neuroscience. Current limitations of Gluc as a reporter include its emission of blue light, which is absorbed by mammalian tissues, limiti...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2013-03, Vol.85 (5), p.3006-3012
Main Authors: Degeling, M. Hannah, Bovenberg, M. Sarah S, Lewandrowski, Grant K, de Gooijer, Mark C, Vleggeert-Lankamp, Carmen L.A, Tannous, Marie, Maguire, Casey A, Tannous, Bakhos A
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Bioluminescence
Cells
Directed Molecular Evolution
Enzymes
Evolution
Immunology
Light
Luciferases - chemistry
Luciferases - genetics
Luciferases - metabolism
Luminescent Measurements
Mammals
Molecular biology
Molecular Sequence Data
Mutation
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Summary:Gaussia Luciferase (Gluc) has proven to be a powerful mammalian cell reporter for monitoring numerous biological processes in immunology, virology, oncology, and neuroscience. Current limitations of Gluc as a reporter include its emission of blue light, which is absorbed by mammalian tissues, limiting its use in vivo, and a flash-type bioluminescence reaction, making it unsuited for high-throughput applications. To overcome these limitations, a library of Gluc variants was generated using directed molecular evolution and screened for relative light output, a shift in emission spectrum, and glow-type light emission kinetics. Several variants with a 10–15 nm shift in their light emission peak were found. Further, a Gluc variant that catalyzes a glow-type bioluminescence reaction, suited for high-throughput applications, was also identified. These results indicate that molecular evolution could be used to modulate Gluc bioluminescence reaction characteristics.
ISSN:0003-2700
1520-6882
DOI:10.1021/ac4003134