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Novel application of Macrolampis sp2 firefly luciferase for intracellular pH-biosensing in mammalian cells

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Abstract

Bioluminescence is widely used in biosensors. Firefly luciferase-based bioluminescent sensors are among the most popular ones. Firefly luciferases are pH-sensitive, displaying a large red shift at acidic pH, a property that has been considered undesirable for most applications. Currently, biosensors that can detect intracellular pH are in demand, and some fluorescent biosensors are available. However, pH sensors using bioluminescence have not been used yet. Thus, we decided to harness a firefly luciferase to measure the intracellular pH in mammalian cells. For this purpose, we engineered the luciferase derived from Macrolampis sp2 firefly to localize it on the cytosol or nucleus, in order to observe pH variation in these compartments during biological activities. We first calibrated the emission ratios (R = Igreen/Ired) at different pH values. As expected, we observed a red shift of light emission under acidic conditions when the cells were subjected to different pH conditions in the presence of the K+/H+ ionophore, nigericin. Based on these results, we concluded that this firefly luciferase can be used as a diagnostic tool for measuring the intracellular pH variation in pathogenic cells or in cells during apoptosis. This is the first example of real time-monitoring of pH change using color tuning luciferase.

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Authors and Affiliations

  1. Graduate School of Biotechnology and Environmental Monitoring, Federal University of São Carlos (UFSCar), Campus of Sorocaba, Sorocaba, São Paulo, Brazil

    Gabriele Verônica de Mello Gabriel & Vadim Ravara Viviani

  2. National Institute of Advanced Industrial Science and Technology (AIST), Biomedical Research Institute, Tsukuba, Japan

    Rie Yasuno & Yoshihiro Ohmiya

  3. National Institute of Advanced Industrial Science and Technology (AIST), Bioproduction Research Institute, Sapporo, Japan

    Yasuo Mitani

  4. Graduate School of Evolutive Genetics and Molecular Biology, Federal University of São Carlos (UFSCar), São Carlos, São Paulo, Brazil

    Vadim Ravara Viviani

Authors
  1. Gabriele Verônica de Mello Gabriel
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  2. Rie Yasuno
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  3. Yasuo Mitani
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  4. Yoshihiro Ohmiya
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  5. Vadim Ravara Viviani
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Correspondence to Vadim Ravara Viviani.

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de Mello Gabriel, G.V., Yasuno, R., Mitani, Y. et al. Novel application of Macrolampis sp2 firefly luciferase for intracellular pH-biosensing in mammalian cells. Photochem Photobiol Sci 18, 1212–1217 (2019). https://doi.org/10.1039/c8pp00573g

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  • DOI: https://doi.org/10.1039/c8pp00573g

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