Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
5131554 | Analytical Biochemistry | 2017 | 4 Pages |
Abstract
Beetle luciferases have been adapted for live cell imaging where bioluminescence is dependent on the cellular availability of ATP, O2, and added luciferin. Previous Photinus pyralis red-emitting variants with high Km values for ATP have performed disappointingly in live cells despite having much higher relative specific activities than enzymes like Click Beetle Red (CBR). We engineered a luciferase variant PLR3 having a Km value for ATP similar to CBR and â¼2.6-fold higher specific activity. The red-emitting PLR3 was â¼2.5-fold brighter than CBR in living HEK293T and HeLa cells, an improvement consistent with the importance of the Km value in low ATP environments.
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Authors
Bruce R. Branchini, Tara L. Southworth, Danielle M. Fontaine, Dawn Kohrt, Franceine S. Welcome, Catherine M. Florentine, Emma R. Henricks, Demetria B. DeBartolo, Elisa Michelini, Luca Cevenini, Aldo Roda, Martha J. Grossel,