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 Kmvalues 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 Kmvalue for ATP similar to CBR and ∼2.6-fold higher specific activity. The red-emitting PLR3 was ∼2.5-fold brighter than CBR in living HEK293T and HeLa cells, an improvement consistent with the importance of the Kmvalue in low ATP environments.

Red-emitting chimeric firefly luciferase for in vivo imaging in low ATP cellular environments

Michelini, Elisa;Cevenini, Luca;Roda, Aldo;
2017

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 Kmvalues 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 Kmvalue for ATP similar to CBR and ∼2.6-fold higher specific activity. The red-emitting PLR3 was ∼2.5-fold brighter than CBR in living HEK293T and HeLa cells, an improvement consistent with the importance of the Kmvalue in low ATP environments.
Branchini, Bruce R.; Southworth, Tara L.; Fontaine, Danielle M.; Kohrt, Dawn; Welcome, Franceine S.; Florentine, Catherine M.; Henricks, Emma R.; Debartolo, Demetria B.; Michelini, Elisa; Cevenini, Luca; Roda, Aldo; Grossel, Martha J.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11585/610204
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