Structure and Function of Activated Rhodopsin

Hofmann, Klaus Peter; Jäger, Stefan; Ernst, Oliver P.

doi:10.1002/ijch.199500035

Cited by 48 publications

(50 citation statements)

References 192 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…MII is the catalytically active intermediate of photoexcited rhodopsin (R*) (1,5,6). We went on to examine whether binding to MII by the two peptides would inhibit Gt activation as expected.…”

Section: Resultsmentioning

confidence: 99%

“…We can now propose that the increasing rate of Gt activation with increasing pH in the acidic branch of the profile reflects the pH dependency of microscopic recognition of domains, which occurs independently of the specific geometry in which they are arranged in conformations MIIa and MIIb, respectively. The decreasing rate at high pH can consistently be attributed (4) to the pH-dependent formation (6,21) of the receptor conformation termed MIIb, which satisfies the grossstructural constraints of catalytic receptor͞G-protein interaction. Although we believe that MIIa is a necessary step in reaching rhodopsin's active conformation (39), its role in nucleotide exchange catalysis (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Metarhodopsin I (MI, max ϭ 478 nm) is in a pH-and temperaturedependent equilibrium with metarhodopsin II (MII, max ϭ 380 nm), distinguished by its deprotonated Schiff base linkage [and broken salt bridge (3,4)] between the retinal and Lys 296 . MII has been shown to catalyze retina rod cell-specific Gprotein (Gt) activation through nucleotide exchange (5,6).…”

mentioning

confidence: 99%

See 2 more Smart Citations

Signal transfer from rhodopsin to the G-protein: Evidence for a two-site sequential fit mechanism

Kisselev

Meyer

Heck

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

105

115

View full text Add to dashboard Cite

Photoactivation of the retinal photoreceptor rhodopsin proceeds through a cascade of intermediates, resulting in protein-protein interactions catalyzing the activation of the G-protein transducin (Gt). Using stabilization and photoregeneration of the receptor's signaling state and Gt activation assays, we provide evidence for a two-site sequential fit mechanism of Gt activation. We show that the C-terminal peptide from the Gt ␥-subunit, Gt␥(50-71)farnesyl, can replace the holoprotein in stabilizing rhodopsin's active intermediate metarhodopsin II (MII). However, the peptide cannot replace the Gt␤␥ complex in direct activation assays. Competition by Gt␥(50-71)farnesyl with Gt for the active receptor suggests a pivotal role for Gt␤␥ in signal transfer from MII to Gt. MII stabilization and competition is also found for the C-terminal peptide from the Gt ␣-subunit, Gt␣(340-350), but the capacity of this peptide to interfere in MII-Gt interactions is paradoxically low compared with its activity to stabilize MII. Besides this disparity, the pH profiles of competition with Gt are characteristically different for the two peptides. We propose a two-site sequential fit model for signal transfer from the activated receptor, R*, to the G-protein.In the center of the model is specific recognition of conformationally distinct sites of R* by Gt␣(340-350) and Gt␥(50-71)farnesyl. One matching pair of domains on the proteins would, on binding, lead to a conformational change in the G-protein and͞or receptor, with subsequent binding of the second pair of domains. This process could be the structural basis for GDP release and the formation of a stable empty site complex that is ready to receive the activating cofactor, GTP.Rhodopsin is a prototypical G-protein-coupled receptor in retinal rods (1, 2). Available information supports a mechanism in which the initial isomerization of the chromophore 11-cis-retinal, and thus the formation of the agonistic all-transretinal, leads to crucial contacts between the ligand and the apoprotein opsin. These steric constraints result in a defined arrangement of donor and acceptor groups for proton translocations leading to subsequent tautomeric conformations of the receptor, identified as ''metarhodopsin'' photointermediates, each with a characteristic absorption spectrum. Metarhodopsin I (MI, max ϭ 478 nm) is in a pH-and temperaturedependent equilibrium with metarhodopsin II (MII, max ϭ 380 nm), distinguished by its deprotonated Schiff base linkage [and broken salt bridge (3, 4)] between the retinal and Lys 296 . MII has been shown to catalyze retina rod cell-specific Gprotein (Gt) activation through nucleotide exchange (5, 6).Despite recent progress in structure determination of both Gt and rhodopsin, the molecular mechanism of signal transfer between the two proteins is poorly understood. Interacting surfaces of rhodopsin and Gt include intracellular loops of the receptor and domains on both Gt ␣-and Gt ␥-subunits (1, 7-9). C-terminal domains of Gt ␣-and Gt ␥-subunits, Gt␣(340-350) and...

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Signal transfer from rhodopsin to the G-protein: Evidence for a two-site sequential fit mechanism

Kisselev

Meyer

Heck

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

105

115

View full text Add to dashboard Cite

show abstract

“…Recently, we found that this structural evolution includes a retinal counterion switch from Glu113 in the dark state to Glu181 in the metarhodopsin I (Meta I) state (19). The Schiff base is then deprotonated upon Meta II formation, and Meta II couples with transducin to trigger the visual signal cascade (20,21).…”

mentioning

confidence: 99%

Resonance Raman Analysis of the Mechanism of Energy Storage and Chromophore Distortion in the Primary Visual Photoproduct

et al. 2004

View full text Add to dashboard Cite

The vibrational structure of the chromophore in the primary photoproduct of vision, bathorhodopsin, is examined to determine the cause of the anomalously decoupled and intense C 11 =C 12 hydrogenout-of-plane (HOOP) wagging modes and their relation to energy storage in the primary photoproduct. Low-temperature (77 K) resonance Raman spectra of Glu181 and Ser186 mutants of bovine rhodopsin reveal only mild mutagenic perturbations of the photoproduct spectrum suggesting that dipolar, electrostatic, or steric interactions with these residues do ded by NIHnot cause the HOOP mode frequencies and intensities. Density functional theory calculations are performed to investigate the effect of geometric distortion on the HOOP coupling. The decoupled HOOP modes can be simulated by imposing ∼40° twists in the same direction about the C 11 =C 12 and C 12 -C 13 bonds. Sequence comparison and examination of the binding site suggests that these distortions are caused by three constraints consisting of an electrostatic anchor between the protonated Schiff base and the Glu113 counterion, as well as steric interactions of the 9-and 13-methyl groups with surrounding residues. This distortion stores light energy that is used to drive the subsequent protein conformational changes that activate rhodopsin.Photoactive proteins play a vital role in a variety of light-energy and light-signaling processes that are characterized by pico-to femtosecond light-driven structural changes of a proteinbound chromophore followed by activated conformational changes of the protein. In systems such as rhodopsin (Rho) 1 (1), bacteriorhodopsin (2,3), halorhodopsin (4), photoactive yellow protein (5), and phytochrome (6,7), primary high-energy intermediates are produced with structurally perturbed chromophores. What is the nature of the protein-chromophore interactions that store the energy needed to drive subsequent protein conformational changes? To address this question, we have studied the photoactivation of rhodopsin, a dim-light photoreceptor, using a multidisciplinary approach that integrates Raman spectroscopy, mutagenesis, density functional theory (DFT) calculations, and bioinformatic analysis. † Funding was provided by NIH Grant EY02051 (to R.A.M. Rho is a paradigmatic G protein-coupled receptor, whose 7-transmembrane helical structure forms a binding pocket for 11-cis-retinal, which is covalently linked to Lys296 on helix 7. Within 200 fs after photoexcitation (8,9), the 11-cis-retinal protonated Schiff base (PSB) isomerizes to all-trans-retinal (ATR) with a quantum yield of 0.65 to form the primary photoproduct, Bathorhodopsin (10,11), which stores ∼30 kcal/mol or ∼60% of the incident photon energy (Figure 1) (12)(13)(14). Subsequently, the highly distorted chromophore relaxes, and the transduced energy is used to drive conformational changes in the opsin protein (15)(16)(17)(18). Recently, we found that this structural evolution includes a retinal counterion switch from Glu113 in the dark state to Glu181 in the metarhodopsin I (Met...

show abstract

“…Absorption of light transforms rhodopsin (rho) into an activated state, which interacts with a heterotrimeric G protein to initiate an enzyme cascade leading to visual transduction. To reach this active state, called metarhodopsin (meta) II (1,2), rho passes through a number of intermediates that have been characterized by UV-visible, Fourier-transform infrared (FTIR), Raman, and NMR spectroscopies, in addition to spin-labeling and biochemical studies (3)(4)(5)(6)(7)(8)(9)(10). One way to characterize the intermediates is to trap them at low temperatures.…”

mentioning

confidence: 99%

Chromophore structural changes in rhodopsin from nanoseconds to microseconds following pigment photolysis

Jäger

Lewis

Zvyaga

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

Rhodopsin is a prototypical G proteincoupled receptor that is activated by photoisomerization of its 11-cis-retinal chromophore. Mutant forms of rhodopsin were prepared in which the carboxylic acid counterion was moved relative to the positively charged chromophore Schiff base. Nanosecond time-resolved laser photolysis measurements of wild-type recombinant rhodopsin and two mutant pigments then were used to determine reaction schemes and spectra of their early photolysis intermediates. These results, together with linear dichroism data, yielded detailed structural information concerning chromophore movements during the first microsecond after photolysis. These chromophore structural changes provide a basis for understanding the relative movement of rhodopsin's transmembrane helices 3 and 6 required for activation of rhodopsin. Thus, early structural changes following isomerization of retinal are linked to the activation of this G protein-coupled receptor. Such rapid structural changes lie at the heart of the pharmacologically important signal transduction mechanisms in a large variety of receptors, which use extrinsic activators, but are impossible to study in receptors using diffusible agonist ligands.Rhodopsin is the light-triggered membrane receptor in rod outer segments responsible for dim-light vision. Absorption of light transforms rhodopsin (rho) into an activated state, which interacts with a heterotrimeric G protein to initiate an enzyme cascade leading to visual transduction. To reach this active state, called metarhodopsin (meta) II (1, 2), rho passes through a number of intermediates that have been characterized by UV-visible, Fourier-transform infrared (FTIR), Raman, and NMR spectroscopies, in addition to spin-labeling and biochemical studies (3-10). One way to characterize the intermediates is to trap them at low temperatures. This approach led to the classical ''bleaching'' scheme ( max values in nanometers): rho (498) 3 bathorhodopsin (batho) (542) 3 lumirhodopsin (lumi) (497) 3 meta I (480) 3 meta II (380) Scheme I However, at more physiologically relevant temperatures, time-resolved measurements up to 1 msec after photolysis reveal a new intermediate and a different kinetic scheme (3, 11). The following reaction scheme was proposed to occur during the nanosecond-to-microsecond time regime (3): rho (498) 3 batho (529) ª bsi (477) 3 lumi (492) 3 . . . Scheme II The new intermediate, blue-shifted intermediate of rho (bsi), is entropically favored and thus not trapped at low temperatures (11).The role of various structural features of the retinal chromophore in early rho photointermediates has been studied through time-resolved spectral measurements of artificial rho, in which the native chromophore is replaced by synthetically modified retinals (12). In studies of bacteriorhodopsin, timeresolved absorption spectroscopy has been shown to be particularly valuable when combined with site-directed mutagenesis to probe the roles of specific protein residues in different intermediates (13). Unti...

show abstract

Structure and Function of Activated Rhodopsin

Cited by 48 publications

References 192 publications

Signal transfer from rhodopsin to the G-protein: Evidence for a two-site sequential fit mechanism

Signal transfer from rhodopsin to the G-protein: Evidence for a two-site sequential fit mechanism

Resonance Raman Analysis of the Mechanism of Energy Storage and Chromophore Distortion in the Primary Visual Photoproduct

Chromophore structural changes in rhodopsin from nanoseconds to microseconds following pigment photolysis

Contact Info

Product

Resources

About