Opsins phosphorylation

At high ambient light (such as noontime outdoors), the level of opsin phosphorylation is such that the protein (3-arrestin binds to the C-terminal segment of opsin. The bound (3-arrestin prevents interaction of Gt with O, totally blocking formation of the active Gta-GTP complex and causing a shutdown of all rod-cell activity. The mechanism by which rod-cell activity is controlled by rhodopsin kinase and ar-restin is similar to adaptation (or desensitization) of other G protein-coupled receptors to high ligand levels. 

FIGURE 127.5 Phosphorylation pattern of opsin. Mass spectra of the rhodopsin AspN cleavage peptide ion from RPE65-deficient mice. Light exposure (A), 24 h dark-adaptation (B), and age do not influence the level or pattern of opsin phosphorylation. 

Ablonczy, Z., Crouch, RK., Goletz, R, Redmond, T.M., Knapp, D.R., Ma, J.-X., and Rohrer, B., 11-cis Retinal reduces constitutive opsin phosphorylation and improves quantum catch in retinoid deficient mouse rod photoreceptors, /. Biol Chem., 277, 40491, 2002. 

S Additional information <1> (<1> RK phosphorylates other color opsins in vivo [13]) [13]... 

S ATP -I- 338-SKTETSQVAPA-348 <1, 12> (<1, 12> peptide containing the last 11 amino acids of the C-terminal of bovine rhodopsin [20, 24] <1> phosphorylated at Ser-343, about 11% of the rate with rhodopsin, photo-activated rhodopsin-dependent, soluble active kinase catalyzes photoacti-vated rhodopsin-independent peptide phosphorylation [20] <12> only in the presence of photoactivated rhodopsin, which activates RK for peptide phosphorylation, also activated by metarhodopsin III, but not by opsin, up to 60% of the rate with photoactivated rhodopsin, light-dependent phosphorylation [24]) (Reversibility <1,12> [20,24]) [20, 24]... 

ATP -I- peptide <1> (<1> monophosphorylated [7] <1> containing sites phosphorylated in rhodopsin [9, 23] <1> less amount of phosphoryl group incorporation than of rhodopsin [9] <1> corresponding to the C-terminus and loop 5-6 of opsin, poor substrates, phosphorylates serine and threonine residues in each peptide [12] <1> acid-rich peptides, RK prefers acid residues localized to the C-terminal side of the serine [15, 23] <1> low catalytic efficiency of RK toward a peptide containing its major autophosphorylation site [27] <1> acidic peptides, stimulated by photo-lyzed rhodopsin, K-491 of RK participates in substrate binding [33]) (Reversibility <1> [7, 9, 12, 15, 22, 23, 27, 33, 36]) [7, 9, 12, 15, 22, 23, 27, 33, 36]... 

The conversion of metarhodopsin II to metarhodopsin III is relatively slow, with a time course of minutes. It is the result of phosphorylation of serine residues in the protein catalyzed by rhodopsin kinase. The final step is hydrolysis to release all-fra s-retinaldehyde and opsin. 

A number of laboratories have also determined that combination of the apoprotein opsin with diW-trans retinal forms a complex which has an activity resembling that of the photolyzed rhodopsin as measured by transducin activation [30], phosphorylation by rhodopsin kinase [31, 32], and arrestin binding [32]. The activity of this complex does not depend on the formation of a Schiff base [33, 34]. The role of such a complex in the physiological process is as yet unclear. 

Mutation of certain amino acids results in rhodopsins that are also constitutively active, that is, the protein can activate transducin in the absence of light [35-37]. Rim and Oprian [38] have shown that rhodopsin mutants that are constitutively active are also phosphorylated by rhodopsin kinase. These results are in agreement with the results on the active opsin complexes discussed above. These mutants disrupt the salt bridge between lysine 296 and glutamate 113 [35-37] and therefore prevent the inactive conformation of the rhodopsin state. These studies have important clinical applications as it has been found that several of the mutations of rhodopsin found in individuals with retinal degenerative disorders result in constitutively active pigments [39]. 

Opsin is the apoprotein of rhodopsin that can result from exposure of the protein to hght (termed bleached opsin) or from lack of the availability of the 11-ds retinal (termed virgin opsin) (Figure 127.4). There is, however, a physiological difference at these two forms of opsin. Bleached opsin is induced by light exposure of rhodopsin and is phosphorylated by rhodopsin kinase. Bleached opsin was shown to have residual activity that reduces visual sensitivity. On the other hand, the characteristics of opsin that has not previously bound 11-c/s retinal, because it was not available due to a disruption in the retinoid metaboHsm " and prior to its developmental occurrence, or because it was unable to bind the Hgand and form rhodopsin, are different from those of the opsin resulting from the photoactivation... 

The MS analysis of the C-terminus of rhodopsin and opsin uncovered unexpected differences between their phosphorylation patterns. These new results may increase our knowledge of how phosphorylation... 

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