EC 1.13.12.24

Reaction: [apoaequorin] + coelenterazine + O₂ + 3 Ca²⁺ = [excited state blue fluorescent protein] + CO₂ (overall reaction)
(1a) [apoaequorin] + coelenterazine = [apoaequorin containing coelenterazine]
(1b) [apoaequorin containing coelenterazine] + O₂ = [aequorin]
(1c) [aequorin] + 3 Ca²⁺ = [aequorin] 1,2-dioxetan-3-one
(1d) [aequorin] 1,2-dioxetan-3-one = [excited state blue fluorescent protein] + CO₂

Glossary: coelenterazine = 8-benzyl-2-(4-hydroxybenzyl)-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one
coelenteramide = N-[3-benzyl-5-(4-hydroxyphenyl)pyrazin-2-yl]-2-(4-hydroxyphenyl)acetamide
aequorin = the non-covalent complex formed by apoaequorin polypeptide and coelenterazine-2-hydroperoxide.
blue fluorescent protein = the non-covalent complex formed by Ca²⁺-bound apoaequorin polypeptide and coelenteramide

Other name(s): Ca²⁺-regulated photoprotein; calcium-activated photoprotein; aequorin; obelin; halistaurin; mitrocomin; phialidin; clytin; mnemiopsin; berovin

Systematic name: coelenterazine:oxygen 2-oxidoreductase (decarboxylating, calcium-dependent)

Comments: Ca²⁺-regulated photoproteins are found in a variety of bioluminescent marine organisms, mostly coelenterates, and are responsible for their light emission. The best studied enzyme is from the jellyfish Aequorea victoria. The enzyme tightly binds the imidazolopyrazinone derivative coelenterazine, which is then peroxidized by oxygen. The hydroperoxide is stably bound until three Ca²⁺ ions bind to the protein, inducing a structural change that results in the formation of a 1,2-dioxetan-3-one ring, followed by decarboxylation and generation of a protein-bound coelenteramide in an excited state. The calcium-bound protein-product complex is known as a blue fluorescent protein. In vivo the energy is transferred to a green fluorescent protein (GFP) by Fšrster resonance energy transfer. In vitro, in the absence of GFP, coelenteramide emits a photon of blue light while returning to its ground state.

Links to other databases: BRENDA, EXPASY, KEGG, Metacyc, CAS registry number:

References:

1. Shimomura, O., Johnson, F. H., and Saiga, Y. Purification and properties of aequorin, a bio-(chemi-) luminescent protein from the jellyfish, Aequorea aequorea. Fed. Proc. 21 (1962) 401.

2. Morise, H., Shimomura, O., Johnson, F.H. and Winant, J. Intermolecular energy transfer in the bioluminescent system of Aequorea. Biochemistry 13 (1974) 2656-2662. [PMID: 4151620]

3. Inouye, S., Noguchi, M., Sakaki, Y., Takagi, Y., Miyata, T., Iwanaga, S., Miyata, T. and Tsuji, F.I. Cloning and sequence analysis of cDNA for the luminescent protein aequorin. Proc. Natl Acad. Sci. USA 82 (1985) 3154-3158. [PMID: 3858813]

4. Head, J.F., Inouye, S., Teranishi, K. and Shimomura, O. The crystal structure of the photoprotein aequorin at 2.3 Å resolution. Nature 405 (2000) 372-376. [PMID: 10830969]

5. Deng, L., Vysotski, E.S., Markova, S.V., Liu, Z.J., Lee, J., Rose, J. and Wang, B.C. All three Ca²⁺-binding loops of photoproteins bind calcium ions: the crystal structures of calcium-loaded apo-aequorin and apo-obelin. Protein Sci. 14 (2005) 663-675. [PMID: 15689515]