title: Measurement of Slow Spontaneous Release of 11-cis-Retinal from Rhodopsin.
abstract: The vertebrate visual photoreceptor rhodopsin (Rho) is a unique G protein-coupled receptor as it utilizes a covalently tethered inverse agonist (11-cis-retinal) as the native ligand. Previously, electrophysiological studies showed that ligand binding of 11-cis-retinal in dark-adapted Rho was essentially irreversible with a half-life estimated to be 420 years, until after thermal isomerization to all-trans-retinal, which then slowly dissociates. This long lifetime of 11-cis-retinal binding was considered to be physiologically important for minimizing background signal (dark noise) of the visual system. However, in vitro biochemical studies on the thermal stability of Rho showed that Rho decays with a half-life on the order of days. In this study, we resolve the discrepancy by measuring the chromophore exchange rate of the bound 11-cis-retinal chromophore with free 9-cis-retinal from Rho in an in vitro phospholipid/detergent bicelle system. We conclude that the thermal decay of Rho primarily proceeds through spontaneous breaking of the covalent linkage between opsin and 11-cis-retinal, which was overlooked in the electrophysiological recording. We estimate that this slow spontaneous release of 11-cis-retinal from Rho should result in 104to 105free opsin molecules in a dark-adapted rod cell-a number that is three orders of magnitude higher than previously expected. We also discuss the physiological implications of these findings on the basal activity of opsins and the associated dark noise in the visual system.
authors.affiliation: Laboratory of Chemical Biology and Signal Transduction, The Rockefeller University, New York, NY.
authors.forename: He
authors.initials: H
authors.lastname: Tian
authors.affiliation: Laboratory of Chemical Biology and Signal Transduction, The Rockefeller University, New York, NY; Department of Neurobiology, Care Sciences and Society, Division of Neurogeriatrics, Center for Alzheimer Research, Karolinska Institutet, Huddinge, Sweden. Electronic address: sakmar@rockefeller.edu.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Sakmar
authors.affiliation: Laboratory of Chemical Biology and Signal Transduction, The Rockefeller University, New York, NY. Electronic address: hubert@rockefeller.edu.
authors.forename: Thomas
authors.initials: T
authors.lastname: Huber
mesh_terms: D003902:Detergents / Q000737:chemistry; D007700:Kinetics; D008823:Micelles; D008958:Models, Molecular; D011487:Protein Conformation; D012172:Retinaldehyde / Q000378:metabolism*; D012243:Rhodopsin / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D003902:Detergents; D008823:Micelles; D012243:Rhodopsin; D012172:Retinaldehyde