Diffusible ligand all-trans-retinal activates opsin via a palmitoylation-dependent mechanism.
In rhodopsin's function as a photoreceptor, 11-cis-retinal is covalently bound to Lys(296) via a protonated Schiff base. 11-cis/all-trans photoisomerization and relaxation through intermediates lead to the metarhodopsin II photoproduct, which couples to transducin (G(t)). Here we have analyzed a different signaling state that arises from noncovalent binding of all-trans-retinal (atr) to the aporeceptor opsin and enhances the very low opsin activity by several orders of magnitude. Like with metarhodopsin II, coupling of G(t) to opsin-atr is sensitive to competition by synthetic peptides from the COOH termini of both G(t)alpha and G(t)gamma. However, atr does not compete with 11-cis-retinal incorporation into the Lys(296) binding site and formation of the light-sensitive pigment. Blue light illumination fails to photorevert opsin-atr to the ground state. Thus noncovalently bound atr has no access to the light-dependent binding site and reaction pathway. Moreover, in contrast to light-dependent signaling, removal of the palmitoyl anchors at Cys(322) and Cys(323) in the rhodopsin COOH terminus impairs the atr-stimulated activity. Repalmitoylation by autoacylation with palmitoyl-coenzyme A restores most of the original activity. We hypothesize that the palmitoyl moieties are part of a second binding pocket for the chromophore, mediating hydrophobic interactions that can activate a large part of the catalytic receptor/G-protein interface.
Sachs
K
K
Institut für Medizinische Physik und Biophysik, Universitätsklinikum Charité, Humboldt-Universität, D-10098 Berlin, Germany.
Maretzki
D
D
Meyer
C K
CK
Hofmann
K P
KP
D000215:Acylation; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D005453:Fluorescence; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D008027:Light; D008969:Molecular Sequence Data; D019308:Palmitic Acid / Q000378:metabolism*; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D011485:Protein Binding; D012172:Retinaldehyde / Q000378:metabolism*; D012243:Rhodopsin / Q000378:metabolism; D012374:Rod Cell Outer Segment / Q000378:metabolism; D017299:Rod Opsins / Q000378:metabolism*; D013053:Spectrophotometry; D015293:Transducin / Q000378:metabolism
D017299:Rod Opsins; D010171:Palmitoyl Coenzyme A; D019308:Palmitic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D012243:Rhodopsin; D015293:Transducin; D012172:Retinaldehyde