Acylation of disc membrane rhodopsin may be nonenzymatic.
Bovine retinal rod outer segments (ROS) support the incorporation of [3H]palmitate into rhodopsin. [14C] Palmitoyl-CoA serves as the donor with an apparent Km of 40 microM. Solubilization of ROS in the detergent, Emulphogene, results in increased incorporation of label into rhodopsin. A further increase is found when ConA-Sepharose-purified rhodopsin is used as the source of both "enzyme" and acceptor. Failure to separate enzyme from acceptor suggested the possibility of a nonenzymatic reaction. This was confirmed when boiled rhodopsin was found to support the reaction. However, the acylation of rhodopsin is not an artifact since analysis of purified native rhodopsin reveals the presence of covalently bound palmitate and we showed that whole bovine retinas incubated with [3H] palmitate incorporated the fatty acid into rhodopsin (O'Brien, P.J., and Zatz, M. (1984) J. Biol. Chem. 259, 5054-5057). Furthermore, in vivo experiments with rat retinas have revealed that opsin is acylated both in the rod inner and outer segments (St. Jules, R. S., and O'Brien, P.J. (1986) Exp. Eye Res. 43, 929-940). Incubation of labeled rhodopsin with mercaptoethanol resulted in release of the labeled palmitate indicating the presence of a thioester bond. This also illustrates the ease with which a thioester, such as palmitoyl cysteine or palmitoyl-CoA, can transfer the fatty acyl group to a free thiol, such as cysteine or mercaptoethanol.
D000215:Acylation; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002417:Cattle; D005227:Fatty Acids / Q000032:analysis; D006358:Hot Temperature; D007700:Kinetics; D008027:Light; D008274:Magnesium / Q000378:metabolism; D015636:Magnesium Chloride; D008623:Mercaptoethanol / Q000494:pharmacology; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D010786:Photoreceptor Cells / Q000378:metabolism*; D012168:Retinal Pigments / Q000378:metabolism*; D012243:Rhodopsin / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000201:enzymology / Q000378:metabolism*
D005227:Fatty Acids; D010169:Palmitic Acids; D012168:Retinal Pigments; D015636:Magnesium Chloride; D010171:Palmitoyl Coenzyme A; D019308:Palmitic Acid; D008623:Mercaptoethanol; D000255:Adenosine Triphosphate; D012243:Rhodopsin; D008274:Magnesium