UC San Diego

Light producing proteins (photoproteins) are attractive molecules to study because of their potential applications in biotechnology and biomedicine. Photoproteins have been identified in several different phyla, however a photoprotein from the phylum Echinodermata has yet to be characterized. The goal in this work is to extract the calcium-dependent photoprotein from brittlestar Ophiopsila californica for purification and identification. The photoprotein was successfully extracted while maintaining activity. Three chromatographic techniques were used to separate the functional photoprotein from other constituents in the brittlestar extracts. The light producing fractions from each of these chromatography methods were analyzed using gel electrophoresis in attempt to identify a band that correlates with the amount of light production when comparing multiple fractions. A ~45 kDa band showed this positive correlation throughout each purification step. This band was characterized by ESI-QTOF-MS but did not result in the identification of a new photoprotein. Similar methods were attempted on isolated photocytes (light producing cells). However nearly all light producing activity was lost when attempting to extract the photoprotein from the cell. This indicated that the photoprotein relies on an intact membrane in order to function. This leads to the idea that the activation of this photoprotein could be different than the one previously proposed.