Transition of Rhodopsin into the Active Metarhodopsin II State Opens a New Light-induced Pathway Linked to Schiff Base Isomerization

Abstract

Rhodopsin bears 11-cis-retinal covalently bound by a protonated Schiff base linkage. 11-cis/all-trans isomerization, induced by absorption of green light, leads to active metarhodopsin II, in which the Schiff base is intact but deprotonated. The subsequent metabolic retinoid cycle starts with Schiff base hydrolysis and release of photolyzed all-trans-retinal from the active site and ends with the uptake of fresh 11-cis-retinal. To probe chromophore-protein interaction in the active state, we have studied the effects of blue light absorption on metarhodopsin II using infrared and time-resolved UVvisible spectroscopy. A light-induced shortcut of the retinoid cycle, as it occurs in other retinal proteins, is not observed. The predominantly formed illumination product contains all-trans-retinal, although the spectra reflect Schiff base reprotonation and protein deactivation. By its kinetics of formation and decay, its low temperature photointermediates, and its interaction with transducin, this illumination product is identified as metarhodopsin III. This species is known to bind alltrans-retinal via a reprotonated Schiff base and forms normally in parallel to retinal release. We find that its generation by light absorption is only achieved when starting from active metarhodopsin II and is not found with any of its precursors, including metarhodopsin I. Based on the finding of others that metarhodopsin III binds retinal in all-trans-C 15 -syn configuration, we can now conclude that light-induced formation of metarhodopsin III operates by Schiff base isomerization ("second switch"). Our reaction model assumes steric hindrance of the retinal polyene chain in the active conformation, thus preventing central double bond isomerization.Living cells react to stimuli, which are realized in physical or chemical signals and are often detected by specialized membrane receptor proteins. G-protein-coupled receptors (GPCRs) 1 transmit their signal to heterotrimeric G-proteins via cytoplasmic domains of their seven-transmembrane ␣-helical structure. The majority of signals are chemical ligands, such as hormones or pheromones, which reach their GPCR by diffusion and bind to a site near their extracellular surface (1, 2). Photoreceptors contain a chromophore as a fixed prostethic group and are specialized for the detection of quanta of visible light in the environment (3, 4). The first step in the signal transduction pathway mediated by these receptors is the generation of a short-lived electronically excited state caused by photon absorption to channel the energy into a conformationally and/or chemically altered state of chromophore-protein interaction (5). Although these early events of light absorption are lacking in ligand binding receptors, G-protein-coupled photoreceptors may use similar mechanisms to spread the local activation near the ligand binding site to the cytoplasmic binding sites, where the G-protein has access. In this view, the chromophore can be understood as a fixed ligand that becomes an agonist by light absorpt...

Evidence for Specificity in Lipid-Rhodopsin Interactions

“…The changes must originate either from Meta-III or opsin. The latter was eliminated by following DHA attenuation over 24 h. The state of rhodopsin with highest magnetization transfer rate toward DHA decayed with a characteristic time of hours as it had been reported for the Meta-III-toopsin conversion (37). The opsin-induced attenuation of DHA was indistinguishable from DHA attenuation by dark-adapted rhodopsin.…”

How a small change in retinal leads to G‐protein activation: Initial events suggested by molecular dynamics calculations

“…32 Other work suggests that both Glu 113 and Glu 181 are unprotonated and that both act as the PSB counterion, with Glu 181 dominating in the Meta I state. 33 We have simulated both Glu 113 and Glu 181 in their unprotonated states, and we find that structural changes occur that could lead to the PSB counterion switching from Glu 113 to Glu 181. The simulation shows that the salt bridge between the retinal PSB and Glu 113 breaks near t = 70 ns after photoisomerization and does not form again (Figs.…”

“…11,32,[54][55][56][57][58][59][60][61][62][63][64][65] This would involve a change in protonation states during the photocycle 33 and experimental work has confirmed that Glu 113 is the dark-state counterion and been suggestive, but not conclusive, that Glu 181 is active during the light-activated stages. 66,67 The simulation results are intriguing in suggesting that this counterion switch mechanism could be present as part of the relaxation mechanism of the protein to the retinal conformational change.…”

The Magnitude of the Light-induced Conformational Change in Different Rhodopsins Correlates with Their Ability to Activate G Proteins

“…Therefore, it is tempting to speculate that during molecular evolution of rhodopsins, vertebrate visual pigments including bovine rhodopsin acquired higher G protein activation ability through acquisition of the larger conformational change, a byproduct of which also resulted in abolishment of photoreversibility in the rhodopsins at the same time. Interestingly, metarhodopsin I, the precursor of the active state of bovine rhodopsin (metarhodopsin II), can be converted back to the dark state by subsequent light absorption (39). Comparison of conformation between metarhodopsin I and the active state of parapinopsin would provide a further evolutionary suggestion.…”