Velamins: the first green-light emitting class of wild-type Ca2+-regulated photoproteins isolated from the ctenophore Velamen parallelum

Soares, D. M.; Galeazzo, G. A.; Sgro, G. G.; Moraes, G. V.; Kronenberg, L.; Borukh, E.; Migotto, A. E.; Gruber, D. F.; Sparks, J. S.; Pieribone, V. A.; Stevani, C. V.; Oliveira, A. G.

doi:10.1101/2024.08.30.610560

bioRxiv · 10.1101/2024.08.30.610560

Velamins: the first green-light emitting class of wild-type Ca2+-regulated photoproteins isolated from the ctenophore Velamen parallelum

Soares, D. M.·Galeazzo, G. A.·Sgro, G. G.·Moraes, G. V.·Kronenberg, L.·Borukh, E.·Migotto, A. E.·Gruber, D. F.·Sparks, J. S.·Pieribone, V. A.·Stevani, C. V.·Oliveira, A. G.

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Abstract

Ca2+-regulated photoproteins (CaPhs) consist of single-chain globular proteins to which coelenterazine, a widely distributed marine luminogenic substrate (the luciferin), binds along with molecular oxygen, producing a stable peroxide. Upon Ca2+ addition, CaPhs undergo conformational changes leading to the cyclization of the peroxide and the formation of a high-energy intermediate. Subsequently, its decomposition yields coelenteramide in an excited state and results in the emission of a flash of light. To date, all CaPhs reported produce blue light ({lambda}max 465-495 nm). Here, we report the cloning and functional characterization of a novel class of wild-type CaPhs capable of emitting green light: velamins, isolated from the bioluminescent ctenophore Velamen parallelum. Ten unique photoprotein-like sequences were recovered and grouped in three main clusters. Representative sequences were cloned, expressed, purified, and regenerated into the active His-tagged -, {beta}-, and {gamma}-velamins. Upon injection of a calcium-containing buffer into the velamin, a flash of green light ({lambda}max 500-508 nm) was observed across pH values ranging from 7 to 9. Whilst -velamin isoforms exhibited the highest light emission activity, {beta}- and {gamma}-velamins were found to be more thermostable at higher temperatures. Velamins are the only known wild-type Ca2+-regulated photoproteins that exhibit the longest wavelength in light emission, making them a promising model for studying spectral modulation. As a result, velamins hold potential for enhancing the sensitivity of signal detection in analytical systems, particularly when dealing with complex biological matrices.

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Soares, D. M., Galeazzo, G. A., Sgro, G. G., Moraes, G. V., Kronenberg, L., Borukh, E., Migotto, A. E., Gruber, D. F., Sparks, J. S., Pieribone, V. A., Stevani, C. V., Oliveira, A. G.. 2024-08-31. Velamins: the first green-light emitting class of wild-type Ca2+-regulated photoproteins isolated from the ctenophore Velamen parallelum. https://doi.org/10.1101/2024.08.30.610560

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