Characterization of rhodopsin kinase in the mammalian retina and pineal gland: A potential prototype for a new class of protein kinases

Rhodopsin kinase, a substrate specific protein kinase found in the rod photoreceptor cells of the retina, phosphorylates the visual pigment rhodopsin subsequent to light absorption. Phosphorylation of photolyzed rhodopsin by rhodopsin kinase decreases its ability to initiate the cascade of reactions leading to visual excitation thereby regulating the sensitivity of the rod photoreceptor cell to light. Rhodopsin kinase was extracted from bovine retinas and purified over 6500-fold utilizing a series of chromatographic steps. Basic to this process was the stabilization of activity with a detergent. Rhodopsin kinase, thus purified, had an apparent molecular weight of 65,000 daltons and underwent adenosine 5$\sp\prime$-triphosphate-dependent autophosphorylation. The enzyme was unstable and subject to proteolysis, initially yielding a 55,000 dalton fragment which was identical in all respects to purified rhodopsin kinase previously reported by Shichi and Somers (1978). Purified rhodopsin kinase is insensitive to regulation by cyclic nucleotides, Ca$\sp{++}$, or lipids. The enzyme is half-maximally inhibited by Ca$\sp{++}$ and Zn$\sp{++}$ at concentrations of 0.1 and 0.5 M, respectively. Additionally, rhodopsin kinase is known to be highly specific for a unique intermediate of rhodopsin photolysis (metarhodopsin II). Rhodopsin kinase is unable to phosphorylate substrates for cyclic-nucleotide-dependent and -independent kinases, as well as substrates for C-kinase and Ca$\sp{++}$/calmodulin-dependent kinase. Rhodopsin kinase activity is highly tissue-specific, being found only in the retina and mammalian pineal organ, a tissue phylogenetically related to the retina. The unique substrate specificity and tissue distribution of the enzyme precludes inclusion of rhodopsin kinase in any of the general classes of protein kinases. The structural and functional similarities between the protein components involved in the light-dependent phosphorylation of rhodopsin and the agonist-dependent phosphorylation of the $\beta$-adrenergic receptor by $\beta$-adrenergic receptor kinase, suggests that the two kinases may exemplify a new class of kinases specific for membrane receptors involved in transmembrane signalling. The present improved method for purification and stabilization of rhodopsin kinase, along with the ability to purify the other components of the light transducing system, make it an ideal model for studying the role of phosphorylation in regulating transmembrane signalling.