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Article Dans Une Revue BBA Advances Année : 2023

Biophysical analysis of Gaussia luciferase bioluminescence mechanisms using a non-oxidizable coelenterazine

Résumé

Gaussia luciferase (GLuc 18.2kDa; 168 residues) is a marine copepod luciferase that emits a bright blue light when oxidizing coelenterazine (CTZ). It is a helical protein where two homologous sequential repeats form two anti-parallel bundles, each made of four helices. We previously identified a hydrophobic cavity as a prime candidate for the catalytic site, but Gluc's fast bioluminescence reaction hampered a detailed analysis. Here, we used azacoelenterazine (Aza-CTZ), a non-oxidizable coelenterazine (CTZ) analog, as a probe to investigate its binding mode to GLuc. Interestingly, the biochemical studies of GLuc inhibition by Aza-CTZ also led us to find that salt, and monovalent anions, are required for GLuc's bioluminescence, which seems reasonable for a sea-dwelling creature. The NMR-based investigation, using chemical shift perturbations monitored by 15NH-HSQC, suggested that Aza-CTZ (and thus unoxidized CTZ) binds to residues in or near the hydrophobic cavity. Of note is that these NMR data are in line with a recent structural prediction of GLuc, hypothesizing that large structural changes occur in regions remote from the hydrophobic cavity upon the addition of CTZ. Interestingly, these results point to- ward a unique mode of catalysis to achieve CTZ oxidative decarboxylation.
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Dates et versions

hal-03978433 , version 1 (10-10-2023)

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Kyoko Takatsu, Naohiro Kobayashi, Nan Wu, Yves Janin, Toshio Yamazaki, et al.. Biophysical analysis of Gaussia luciferase bioluminescence mechanisms using a non-oxidizable coelenterazine. BBA Advances, 2023, 3, pp.100068. ⟨10.1016/j.bbadva.2022.100068⟩. ⟨hal-03978433⟩
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