Reducing agents and light break an s-s bond activating rhodopsin in vivo in Chlamydomonas

Abstract

Light induces retinal synthesis via photoactivation of a small amount of Chlamydomonas rhodopsin pigment (Foster et al., Proc. Natl. Acad. Sci. USA 85, 6379-6383). A reducing agent [dithiothreitol (DDT) or mercaptoacetic acid (MAA)] also induces retinal synthesis in the dark via a rhodopsin with a chromophore. If the opsin is saturated with retinal and is bleached with light in the presence of a thiol trapping agent, the bleaching becomes irreversible. We conclude that the reducing agent as well as light break a disulfide bond resulting in activation of the rhodopsin and induction of carotenogenesis. Both the chemical and light induction is inhibited by GDPβS and pertussis toxin. Breaking the bridge between the 3rd and 5th helix may lead to increased proton accessibility of Asp134 leading to the rolling of the 3rd helix and MetaIIb formation. (C) 2000 Academic Press.