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

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 ?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 Km value in low ATP environments.

SUBMITTER: Branchini BR

PROVIDER: S-EPMC5730985 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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

Branchini Bruce R BR Southworth Tara L TL Fontaine Danielle M DM Kohrt Dawn D Welcome Franceine S FS Florentine Catherine M CM Henricks Emma R ER DeBartolo Demetria B DB Michelini Elisa E Cevenini Luca L Roda Aldo A Grossel Martha J MJ

Analytical biochemistry 20170704

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 ∼2.6-fo ...[more]

PMID: 28687486

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