Calcium ion is a ubiquitous intracellular messenger, performing this function in many eukaryotic cells. To understand calcium regulation mechanisms and how disturbances of these mechanisms are associated with disease states, it is necessary to measure calcium inside cells. Ca
2+-regulated photoproteins have been successfully used for this purpose for many years. Here we report the results of comparative studies on the properties of recombinant aequorin from
Aequorea victoria, recombinant obelins from
Obelia geniculata and
Obelia longissima, recombinant mitrocomin from
Mitrocoma cellularia, and recombinant clytin from
Clytia gregaria as intracellular calcium indicators in a set of identical in vitro and in vivo experiments. Although photoproteins reveal a high degree of identity of amino acid sequences and spatial structures, and, apparently, have a common mechanism for the bioluminescence reaction, they were found to differ in the Ca
2+ concentration detection limit, the sensitivity of bioluminescence to Mg
2+, and the rates of the rise of the luminescence signal with a sudden change of Ca
2+ concentration. In addition, the bioluminescence activities of Chinese hamster ovary cells expressing wild-type photoproteins also differed. The light signals of cells expressing mitrocomin, for example, slightly exceeded the background, suggesting that mitrocomin may be hardly used to detect intracellular Ca
2+ without modifications improving its properties. On the basis of experiments on the activation of endogenous P2Y
2 receptor in Chinese hamster ovary cells by ATP, we suggest that wild-type aequorin and obelin from
O. longissima are more suitable for calcium detection in cytoplasm, whereas clytin and obelin from
O. geniculata can be used for calcium measurement in cell compartments with high Ca
2+ concentration.
Figure Hydromedusan photoproteins differ in Ca2+ concentration detection limit, sensitivity of bioluminescence to Mg2+, and rates of rise of luminescence signal with a sudden change of [Ca2+] despite a high degree of identity of their amino acid sequences and spatial structures, and, apparently, a common mechanism for the bioluminescence reaction. 相似文献