[16] Use of α-toxin-permeabilized photoreceptors in in vitro phototransduction studies

Publisher Summary In vitro assays utilizing purified retinal proteins have been a cornerstone of biochemical studies of phototransduction. However, many studies have relied on purification and assay techniques that destroy photoreceptor integrity, and consequently, require the evaluation of dynamic enzymatic interactions under nonphysiological conditions. In vitro study of phototransduction, using intact photoreceptors, is an appealing alternative to conventional biochemical techniques. This strategy preserves conditions for enzymatic interactions that more closely resemble those found in vivo . A variety of permeabilization strategies have been developed to overcome the physical barrier imposed by the plasma membrane. One of these techniques, toxin-mediated poration, has been used in a growing number of in vitro studies in both whole cells and membranebound organelles. This technique was valuable in our own studies of rhodopsin phosphorylation in intact photoreceptors. The utility of the α-toxin as a permeabilizing agent has been substantially enhanced through the engineering of gateable pores composed of mutant α-toxin proteins.