Properties of the Chaetopterus Photoprotein and its Luminescence Reaction

Spectral properties. The purified photoprotein is practically colorless, although its absorption spectrum (Fig. 7.1.4) shows a very slight absorption in the region of 330-380nm in addition to the 280 nm protein peak. The solution of photoprotein is moderately blue fluorescent, with an emission maximum at 453-455 nm and an 

Luminescence reaction. In air, partially purified preparations of the photoprotein emit light in the presence of Fe2+ and a peroxide however, highly purified preparations require two additional substances, cofactors 1 and 2, to emit the same intensity of light. Both cofactors can be substituted with a small amount of a crude extract of the photoprotein. 

To measure the total light emission of a photoprotein sample, it was necessary to add 1-5 pM Fe2+ several times due to the short effective life of Fe2+ under the conditions involved. The total amount of light measured in this manner was always proportional to the weight of photoprotein used, with both CPA and CPC (4.7 x 1013 photons/mg at 25°C Shimomura and Johnson, 1968d). Thus, the quantum yield is estimated at roughly 0.01 for CPA, and 0.015 for CPC. 

No information is available on the chemical nature of the luminophore, although the photoprotein must contain a chro-mophore to emit luminescence and fluorescence. Acid treatment of the protein, followed by extraction with organic solvents, did not yield coelenteramide or coelenteramine, indicating that this luminescence system is unrelated to coelenterazine. A flavin (FAD) was found in partially purified preparations of photoprotein, but not in highly purified preparations. 

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