All Ca2 +-binding loops of light-sensitive ctenophore photoprotein berovin bind magnesium ions: The spatial structure of Mg2 +-loaded apo-berovin

• The spatial structure of apo-berovin with bound Mg2 + is determined at 1.75 Å.
• Magnesium ions are found in each functional EF-hand loop of a photoprotein.
• Mg2 + is coordinated by oxygen atoms with characteristic oxygen-Mg2 + distances.
• These findings account for the influence of Mg2 + on berovin bioluminescence.

Light-sensitive photoprotein berovin accounts for a bright bioluminescence of ctenophore Beroe abyssicola. Berovin is functionally identical to the well-studied Ca2 +-regulated photoproteins of jellyfish, however in contrast to those it is extremely sensitive to the visible light. Berovin contains three EF-hand Ca2 +-binding sites and consequently belongs to a large family of the EF-hand Ca2 +-binding proteins. Here we report the spatial structure of apo-berovin with bound Mg2 + determined at 1.75 Å. The magnesium ion is found in each functional EF-hand loop of a photoprotein and coordinated by oxygen atoms donated by the side-chain groups of aspartate, carbonyl groups of the peptide backbone, or hydroxyl group of serine with characteristic oxygen-Mg2 + distances. As oxygen supplied by the side-chain of the twelfth residue of all Ca2 +-binding loops participates in the magnesium ion coordination, it was suggested that Ca2 +-binding loops of berovin belong to the mixed Ca2 +/Mg2 + rather than Ca2 +-specific type. In addition, we report an effect of physiological concentration of Mg2 + on bioluminescence of berovin (sensitivity to Ca2 +, rapid-mixed kinetics, light-sensitivity, thermostability, and apo-berovin conversion into active protein). The different impact of physiological concentration of Mg2 + on berovin bioluminescence as compared to hydromedusan photoproteins was attributed to different affinities of the Ca2 +-binding sites of these photoproteins to Mg2 +.

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