Use of Photoproteins as Intracellular Calcium Indicators

The calcium-regulated photoproteins, of which aequorin is the best known, continue to be one of the most useful groups of intracellular Ca2+indicators. They are self-contained bioluminescent systems that emit blue light in the presence of Ca2+ions, can readily be purified intact, and are nontoxic when introduced into foreign cells. They have been used successfully as Ca2+indicators in almost every kind of cell, but are most widely used in muscle cells because of their relative freedom from motion artifacts. Photoproteins have also been used in conjunction with microscopic image intensification to localize Ca2+in cells. Their large molecular size makes them difficult to introduce into cells, but once there, they have the advantage of staying in the cytoplasm. Aequorin can be microinjected satisfactorily into single cells of almost any size, but a number of alternative methods for introducing photoproteins into cells have been developed in recent years. Disadvantages of the photoproteins for some applications include the nonlinear relation between [ Ca2+] and light intensity, the modest speed with which they respond to sudden changes in [ Ca2+], and the fact the Mg2+antagonizes the effect of Ca2+. Native photoproteins consist of a mixture of isospecies, and there are differences in Ca2+sensitivity and in kinetic properties-both among photoproteins and among the isospecies of a given photoprotein. The genes for several of the isospecies of aequorin have been cloned and expressed in E. coli. It seems reasonable to hope that genetic engineering techniques may soon make it possible to consider using, as Ca2+indicators, rare isospecies or rare photoproteins that have optimal properties for particular applications.