INHIBITION OF RHODOPSIN KINASE BY RECOVERIN - FURTHER EVIDENCE FOR A NEGATIVE FEEDBACK-SYSTEM IN PHOTOTRANSDUCTION

Recoverin is a 23-kDa Ca2+-binding protein found predominantly in vert ebrate photoreceptor cells. Recent electrophysiological and biochemica l studies suggest that recoverin may regulate the photoresponse by inh ibiting rhodopsin phosphorylation. We find in both cell homogenates an d reconstituted systems that the inhibition of rhodopsin phosphorylati on by recoverin occurs over a significantly higher free Ca2+ range tha n previously reported, Half-maximal inhibition occurs at 1.5-3 mu M fr ee Ca2+ and is cooperative with a Hill coefficient of similar to 2. Me asurements of transducin activation demonstrate that this inhibition p rolongs the lifetime of catalytically active rhodopsin. Ca2+-recoverin directly inhibits rhodopsin kinase activity, and Ca2+-dependent bindi ng of recoverin to rod outer segment membranes is not required for its action, Extrapolation of the in vitro data to in vivo conditions base d on simple mass action calculations places the Ca2+-recoverin regulat ion within the physiological free Ca2+ range in intact rod outer segme nt. The data are consistent with a model in which the fall in free Ca2 + that accompanies rod excitation exerts negative feedback by relievin g inhibition of rhodopsin phosphorylation.