A catalytic mechanism for Renilla-type bioluminescence

Schenkmayerova, A.; Toul, M.; Pluskal, D.; Baatallah, R.; Gagnot, G.; Pinto, G. P.; Santana, V. T.; Stuchla, M.; Neugebauer, P.; Chaiyen, P.; Damborsky, J.; Bednar, D.; Janin, Y. L.; Prokop, Z.; Marek, M.

doi:10.1101/2022.02.09.479090

bioRxiv · 10.1101/2022.02.09.479090

A catalytic mechanism for Renilla-type bioluminescence

Schenkmayerova, A.·Toul, M.·Pluskal, D.·Baatallah, R.·Gagnot, G.·Pinto, G. P.·Santana, V. T.·Stuchla, M.·Neugebauer, P.·Chaiyen, P.·Damborsky, J.·Bednar, D.·Janin, Y. L.·Prokop, Z.·Marek, M.

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Abstract

The widely used coelenterazine-powered Renilla luciferase was discovered over 40 years ago but the oxidative mechanism by which it generates blue photons remains unclear. Here we decipher Renilla-type bioluminescence through crystallographic, spectroscopic, and computational experiments. Structures of ancestral and extant luciferases complexed with the substrate-like analogue azacoelenterazine or a reaction product were obtained, providing unprecedented snapshots of coelenterazine-to-coelenteramide oxidation. Bound coelenterazine adopts a Y-shaped conformation, enabling the deprotonated imidazopyrazinone component to attack O2 via a radical charge-transfer mechanism. A high emission intensity is secured by an aspartate from a conserved proton-relay system, which protonates the excited coelenteramide product. Another aspartate on the rim of the catalytic pocket fine-tunes the electronic state of coelenteramide and promotes the formation of the blue light-emitting phenolate anion. The results obtained also reveal structural features distinguishing flash-type from glow-type bioluminescence, providing insights that will guide the engineering of next-generation luciferase-luciferin pairs for ultrasensitive optical bioassays.

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Schenkmayerova, A., Toul, M., Pluskal, D., Baatallah, R., Gagnot, G., Pinto, G. P., Santana, V. T., Stuchla, M., Neugebauer, P., Chaiyen, P., Damborsky, J., Bednar, D., Janin, Y. L., Prokop, Z., Marek, M.. 2022-02-09. A catalytic mechanism for Renilla-type bioluminescence. https://doi.org/10.1101/2022.02.09.479090

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