Preparation and X-ray crystallographic analysis of the Ca2+-discharged photoprotein obelin

Ca(2+)-regulated photoproteins belong to the EF-hand Ca(2+)-binding protein family. The addition of calcium ions initiates bright blue bioluminescence of the photoproteins, a result of the oxidative breakdown of coelenterazine peroxide to coelenteramide. Crystals of the Ca(2+)-discharged W92F mutant of obelin from Obelia longissima have been grown, representing the first crystallization of a photoprotein after the Ca(2+)-triggered bioluminescence. A green fluorescence observed from the crystals clearly demonstrates that coelenteramide, the bioluminescence product of coelenterazine peroxide, is bound within the protein. The diffraction pattern exhibits tetragonal Laue symmetry. Systematic absences indicate that the space group is either P4(3)2(1)2 or P4(1)2(1)2. The unit-cell parameters are a = b = 53.4, c = 144.0 A. The crystals diffract to 1.9 A resolution.