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 vertebrate photoreceptor cells. Recent electrophysiological and biochemical studies suggest that recoverin may regulate the photoresponse by inhibiting rhodopsin phosphorylation. We find in both cell homogenates and reconstituted systems that the inhibition of rhodopsin phosphorylation by recoverin occurs over a significantly higher free Ca2+ range than previously reported, Half-maximal inhibition occurs at 1.5-3 mu M free Ca2+ and is cooperative with a Hill coefficient of similar to 2. Measurements of transducin activation demonstrate that this inhibition prolongs the lifetime of catalytically active rhodopsin. Ca2+-recoverin directly inhibits rhodopsin kinase activity, and Ca2+-dependent binding of recoverin to rod outer segment membranes is not required for its action, Extrapolation of the in vitro data to in vivo conditions based on simple mass action calculations places the Ca2+-recoverin regulation within the physiological free Ca2+ range in intact rod outer segment. The data are consistent with a model in which the fall in free Ca2+ that accompanies rod excitation exerts negative feedback by relieving inhibition of rhodopsin phosphorylation.