Protein kinases of retinal rod outer segments: identification and partial characterization of cyclic nucleotide dependent protein kinase and rhodopsin kinase.
Protein kinase activity of dark-adapted bovine rod outer segments is partitioned by centrifugation into soluble and membrane-bound fractions. The soluble kinases are separated by DEAE-cellulose chromatography into three peaks of activity, which can be classified by substrate specificity and cyclic nucleotide dependence into two categories. One peak of protein kinase activity has the characteristics reported for rhodopsin kinase (category one); it phosphorylates only bleached rhodopsin, and its activity is not affected by light, exogenous adenosine cyclic 3',5'--monophosphate (cAMP), guanosine cyclic 3',5'-monophosphate (cGMP), or a protein kinase inhibitor from skeletal muscle. Rhodopsin kinase has an apparent molecular weight of 68 000. The second category of kinase includes two peaks of activity which are stimulated severalfold by cAMP or cGMP but not by light. These protein kinases phosphorylate soluble proteins including histones and a protein kinase substrate prepared from rat intestine but not rhodopsin. The two peaks elute from DEAE-cellulose with 0.09 and 0.20 M KCl, suggesting that they are similar respectively to type I and type II cyclic nucleotide dependent protein kinases that have been characterized in other tissues. The activity of type I kinase is variable and much less than that of the type II enzyme; its molecular weight was not determined. The type II protein kinase has an apparent molecular weight of 165 000. This study confirms that different protein kinase enzymes catalyze selectively the phosphorylation of bleached rhodopsin and soluble proteins, and it repudiates the speculation in a previous publication [Farber, D. B., Brown, B. M., & Lolley, R. N. (1979) Biochemistry 18, 370-378] that a single protein kinase might catalyze both phosphorylation reactions.
D000818:Animals; D002417:Cattle; D000242:Cyclic AMP / Q000494:pharmacology*; D006152:Cyclic GMP / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D005136:Eye Proteins; D051606:G-Protein-Coupled Receptor Kinase 1; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D010786:Photoreceptor Cells / Q000201:enzymology*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000302:isolation & purification*; D011817:Rabbits; D051381:Rats; D012243:Rhodopsin / Q000302:isolation & purification; D012374:Rod Cell Outer Segment / Q000201:enzymology*; D012995:Solubility; D013379:Substrate Specificity
D005136:Eye Proteins; D047428:Protein Kinase Inhibitors; D012243:Rhodopsin; D000242:Cyclic AMP; D011494:Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1; C496765:Grk1 protein, rat; D006152:Cyclic GMP