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Influence of the C-terminal domain on the bioluminescence activity and color determination in green and red emitting beetle luciferases and luciferase-like enzyme

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Abstract

Beetle luciferases catalyze the bioluminescent oxidation of D-luciferin, producing bioluminescence colors ranging from green to red, using two catalytic steps: adenylation of D-luciferin to produce D-luciferyl-adenylate and PPi, and oxidation of D-luciferyl-adenylate, yielding AMP, CO2, and excited oxyluciferin, the emitter. Luciferases and CoA-ligases display a similar fold, with a large N-terminal domain, and a small C-terminal domain which undergoes rotation, closing the active site and promoting both adenylation and oxidative reactions. The effect of C-terminal domain deletion was already investigated for Photinus pyralis firefly luciferase, resulting in a red-emitting mutant with severely impacted luminescence activity. However, the contribution of C-terminal in the bioluminescence activities and colors of other beetle luciferases and related ancestral luciferases were not investigated yet. Here we compared the effects of the C-terminal domain deletion on green-emitting luciferases of Pyrearinus termitilluminans (Pte) click beetle and Phrixothrix vivianii railroadworm, and on the red-emitting luciferase of Phrixothrix hirtus railroadworm and luciferase-like enzyme of Zophobas morio. In all cases, the domain deletion severely impacted the overall bioluminescence activities and, slightly less, the oxidative activities, and usually red-shifted the bioluminescence colors. The results support the involvement of the C-terminal in shielding the active site from the solvent during the light emitting step. However, in Pte luciferase, the deletion caused only a 10 nm red-shift, indicating a distinctive active site which remains more shielded, independently of the C′-terminal. Altogether, the results confirm the main contribution of the C-terminal for the catalysis of the adenylation reaction and for active site shielding during the light emitting step.

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Acknowledgements

FAPESP 2010/05426-8, CNPq 401867/2016-1.

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Authors and Affiliations

  1. Graduate Program of Evolutive Genetics and Molecular Biology, Federal University of São Carlos (UFSCar), São Carlos, Brazil

    V. R. Bevilaqua, Mariele C. Carvalho & Vadim R. Viviani

  2. Department of Physics, Chemistry and Mathematics, Federal University of São Carlos (UFSCar), Sorocaba, Brazil

    Gabriel F. Pelentir & Vadim R. Viviani

  3. Brazilian Biorenewables National Laboratory (LNBR), Brazilian Center for Research in Energy and Materials (CNPEM), Campinas, SP, Brazil

    Atílio Tomazini & Mario Murakami

  4. Graduate Program of Biotechnology and Environmental Monitoring, Federal University of São Carlos (UFSCar), Rod. João Leme Dos Santos, SP 264, Km 110, Itinga, Sorocaba, SP, Brazil

    Vadim R. Viviani

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  1. V. R. Bevilaqua
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  2. Mariele C. Carvalho
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  4. Atílio Tomazini
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Contributions

VB produced Re and Pte deletion mutants, MC produced and characterized Zop deletion mutant, GFP characterized Pte and RE deletion mutants, AT made and discussed the three-dimensional models, VV idealized the work, prepared the manuscript and produced pxGR deletion mutant.

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Correspondence to Vadim R. Viviani.

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Bevilaqua, V.R., Carvalho, M.C., Pelentir, G.F. et al. Influence of the C-terminal domain on the bioluminescence activity and color determination in green and red emitting beetle luciferases and luciferase-like enzyme. Photochem Photobiol Sci 20, 113–122 (2021). https://doi.org/10.1007/s43630-020-00007-5

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  • DOI: https://doi.org/10.1007/s43630-020-00007-5

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