title: Rhodopsin phosphorylation occurs at metarhodopsin II level.
abstract: Photolyzed rhodopsin was phosphorylated in bovine rod outer segments incubated at -10 degrees C. In the experiment in which urea-treated outer segments and rhodopsin kinase were incubated with ATP in the presence of 30% glycerol, the extent of phosphate incorporation at -10 degrees C was about 30% of that at 37 degrees C. Separation of phosphorylated rhodopsin by isoelectric focusing indicated that a limited number of sites were phosphorylated at -10 degrees C. The partially phosphorylated pigment incorporated more phosphates when the temperatures was raised to 37 degrees C. This was partly due to decreased inhibition of phosphorylation by glycerol at higher temperature. Since the maximum phosphorylation at -10 degrees C (at which metarhodopsin II is stable) occurred at a pH value (6.0) lower than the pKa for metarhodopsin I-metarhodopsin II transition, metarhodopsin II was suggested to be the preferred substrate for rhodopsin kinase at -10 degrees C. Limited proteolysis with thermolysin of rhodopsin phosphorylated at 37 degrees C released peptides containing about 50% of the total phosphate incorporated. In contrast, proteolytic digestion of rhodopsin phosphorylated at -10 degrees C released negligible amounts of phosphate-containing peptides. The results were taken to suggest that the incorporation of phosphates at metarhodopsin II level under the present condition occurred in the residues other than those removed by thermolysin digestion.
mesh_terms: D000818:Animals; D002417:Cattle; D005136:Eye Proteins; D051606:G-Protein-Coupled Receptor Kinase 1; D007700:Kinetics; D010766:Phosphorylation; D010786:Photoreceptor Cells / Q000201:enzymology*; D011494:Protein Kinases / Q000378:metabolism*; D012168:Retinal Pigments / Q000378:metabolism*; D012243:Rhodopsin / Q000031:analogs & derivatives / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000201:enzymology*
chemical_list: D005136:Eye Proteins; D012168:Retinal Pigments; C013110:metarhodopsins; D012243:Rhodopsin; D011494:Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1