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===Light-Induced Visual Signal Transduction===
===Light-Induced Visual Signal Transduction===
-
Light absorption and G protein activation
+
In the active site of rhodopsin, there is a protonated 11-cis retinal (retinylidene) Schiff base which functions as an inverse agonist.
 +
In photoisomeration, the primary step in rhodopsin photoactivation, 11-cis retinylidene ligand is switched into an all-trans retinal configuration. In this <200 fs process, the protein-binding pocket accommodates the 11-cis conformation of the chromophore and is preserved, restraining the relaxation of the chromophore. The strained chromophore relaxation of conformational energy transforms the protein from the inactive to active form<ref>Article 2</ref>.
 +
 
 +
Rhodopsin is converted into the active signalling state of metarhodopsin MII.
 +
Rhodopsin kinase phosphorylates rhodopsin and arrestin binds to rhodopsin, preventing further signal transduction from MII.
 +
The all-trans¬-retinal Schiff base linkage of MII may be hydrolyzed and cleaved, releasing free retinal and leaving the apoprotein, opsin.
 +
Free retinal can be enzymatically converted into 11-cis-retinal form, then rebinds to opsin to regenerate rhodopsin<ref>Article 3</ref>
 +
When metarhodopsin II is formed, Schiff base deprotonates and proton is transferred to Glu113, releases stored light energy and activates the protein.
 +
The retinal protonated Schiff base and Glu113 efficiently locks the receptor in an inactive state in the dark.
 +
 
 +
Rhodopsin can convert light energy into a chemical signal immensely efficiently.<ref>Article 4</ref>
 +
 
 +
Signal transduced is a “ligand” consisting of a photon interacting with a covalently bound retinal, changing the conformation of the retinal and opsin (protein moiety). This generates a signalling cascade where a large number of transducin molecules, the trimeric G protein in the cytoplasm, are activated through trigger of GDP-GTP nucleotide exchange in the α subunit. This results in the hyperpolarisation of the rod photoreceptor cells and synaptic signalling to adjacent rod bipolar cells; each activated transducin activates cGMP-specific phosphodiesterase, which results in the hydrolysis of cGMP molecules and the closing of the cGMP-gated cation channels in the plasma membrane of the outer segment. The cell hyperpolarises due to the decrease in the influx of sodium and calcium ions, results in the decrease in the release of glutamate into the synaptic cleft.
 +
....For the post-synaptic state, the reduced glutamate levels inhibit G protein-mediated signalling by mGluR6 receptors resulting in the opening of the cation channels, depolarisation, and transmittance of the signals from the retina through nerve impulses to the visual system.
 +
11-cis-retinal seems to act as an inverse agonist, repressing opsin activity, stabilizing the structural inactive conformation.
 +
When activated by light, a photochemical reaction occurs in picoseconds where the retinal is converted into an all-trans form. Following that is a slower thermal relaxation process involving retinal and opsin undergoing conformational changes to result in fully active metarhodopsin II. <ref>Article 6</ref>.
 +
 
 +
It phosphorylates both metarhodopsin I and II and cone opsins. The majority of the phosphorylation sites were in the cytoplasmic C-terminal region of rhodopsin with seven hydroxy-amino acids. Most favoured were Ser 338, Ser 343, Ser 334, Thr 335 and Thr 336.
 +
Protein phosphatise 2A recognizes and dephosphorylates rhodopsin.<ref>Article 7</ref>.
 +
 
 +
 
 +
The 11-cis-retinal is permanently bound to the receptor as a prosthetic group and inactivates it.
 +
The retinal plays a chromophore in the initial photochemistry state and as an agonist after the relaxation state in producing the active state of the receptor.
 +
To desensitize signalling to a G protein, phosphorylating the activated receptor is one method. The major phosphorylation sites of rhodopsin are Ser 338, Ser 343 and Ser 334. These residues form an arrangement that does not appear to be exposed to the solvent. Interactions with the C-terminal tail and a portion of the C-III loop appear to be broken for the phosphorylation of the hydroxyl groups.
 +
1) Light induced isomerization of the retinal from cis to trans. Fits well in retinal pocket.
 +
2) thermal relaxation of the retinal-protein complex
 +
3) Late equilibria affected by rhodopsin and G protein interaction.
 +
The meta states:
 +
Metarhodopsin I forms to Metarhodopsin II which is the intermediate signalling state where interaction occurs with the G protein.
 +
This is accompanied by the breakage of the salt bridge between Glu 113 and the protonated Schiff base between Lys 296 and retinal, and movement in the helices. Occurs in milliseconds. The Schiff base proton is transferred to the Glu 113 counterion., thus destabilizing the ground state.
 +
The cytoplasm also uptakes protons in the formation of MII. Is speculated to be dependent on the protonation of Glu134, a highly conserved residue, which destabilizes the constraint on the Arg 135 it is salt-bridged with.
 +
In forming the signalling state, there is positive enthalpy with the formation of MII. Formation of the active state, linked with the increase in entropy, is suggested to release the constrains in the helices and expose the cytoplasmic binding sites.
 +
The 9-methyl group of retinal is suggested to provide a scaffold for proton transfers essential for the formation of the active state<ref>Article 9</ref>.
 +
 
 +
Like most GCPRs though, the activated rhodopsin catalyzes uptake of GTP by heterotrimeric G protein, in this case transducin, which interacts with cytoplasmic loops of the receptor. Dissociation of the GTP-bound α-subunit from the βγ heterodimer enables triggering of the rod cell’s response to light.
 +
Upon activation, movement and slight adjustment of helices are observed, with the inner faces of Helix II, III, VI and VII more exposed<ref>Article 10</ref>.
 +
 
 +
Different states including short-lived, photo- batho- and lumi-rhodopsin, and longer-lived meta-rhodopsins give information of the structural status of the molecule during activation.
 +
Upon activation, Meta II state formation is accompanied by deprotonation of the Schii base, breakage of the salt bridge with couterion residue Glu 113.
 +
Upon activation, helices three and six move outwards, binding site for transducin is accessible as opening between cytoplasmic loops.
 +
Deactivation occurs when rhodopsin kinase phosphorylates rhodopsin’s C-terminal region, and arrestin binds, competing with transducin. Proceeds by decay of Meta II to opsin. For activation of rhodopsin, rhodopsin has to be dephosphorylated, and have the 11-cis retinal form again<ref>Article 19</ref>.
 +
 
==Opsin==
==Opsin==

Revision as of 06:12, 26 March 2010

PDB ID 1u19

Drag the structure with the mouse to rotate
1u19, resolution 2.20Å ()
Ligands: , , , , , , , ,
Non-Standard Residues:
Related: 1f88, 1hzx, 1l9h
Resources: FirstGlance, OCA, PDBsum, RCSB
Coordinates: save as pdb, mmCIF, xml


Contents

Introduction

Rhodopsin

Rhodopsin, a dimeric protein, is a highly characterized G protein-coupled receptor found in the neurons of the retina and in rod photoreceptor cells. It is part of the superfamily of G protein-coupled receptors that mediate responses to visual, olfactory, hormonal, and neurotransmitter signals among others[1]. Rhodopsin is known as the visual pigment, comprising of an opsin apoprotein and an 11-cis retinal chromophore linked to Lysine 296 by a protonated Schiff base[2].

G Protein-Coupled Receptors

Rhodopsin is a member of the superfamily of G protein-coupled receptors which incorporate the activation of G proteins in their mediation of signalling and intracellular actions. Rhodopsin shares similar membrane topology with the members of the superfamily (Family A of the G protein-coupled receptors) which include the seven transmembrane helices, an extracellular N terminus and cytoplasmic C terminus[3]. As the crystal structure for any G protein-coupled receptor with the seven transmembrane domain has only been solved for rhodopsin, rhodopsin may be a reference for the structure and function relationship for other G protein-coupled receptors[4]. However, the covalent binding nature of rhodopsin to its retinal ligand is unlike most G protein-coupled receptors. As well, another difference of rhodopsin from the members of this superfamily relates to light as the inducer for activation[5].

Structure

Rhodopsin Architecture

Rhodopsin consists of seven mostly α-helical transmembrane domains (H1-H7) linked sequentially by extracellular and cytoplasmic loops (E1-E3 and C1-C3 respectively), with the extracellular amino-terminal tail and the cytoplasmic carboxyl-terminal tail[6]. Four of the helices are tilted and three of the helices are approximately perpendicular to the membrane plane[7]. There is notable interaction between the four extracellular domains, but only a few associations are observed with the cytoplasmic domains[8].

Also, there is the presence of a cationic amphipathic Helix 8, known as the fourth cytoplasmic loop, that is formed from the C-terminal tail anchoring to the membrane by two cysteines, which include palmitates in the structure. This helix runs approximately parallel to the cytoplasmic surface and is involved in Gtγ binding[9], as well as the modulation of rhodopsin-transducin interactions and rhodopsin-phospholipid interactions[10].

Helix 7 is close to being elongated around the Lysine 296 retinal attachment site, and also contains the residues Proline 291 and Proline 303, with Proline 303 being part of a conserved motif[11]. Near the retinal region, there is a β4 strand within the Extracellular Helix 2 that runs almost parallel to the chromophore held in place, and is stabilized by the essential conserved disulfide bond between Cysteine 110 and Cysteine 187[12]. The protonated Schiff base is stabilized through Glutamine 113 residue electrostatic ineraction with the counterion, holding the inactive rhodopsin in its state[13]. A metal zinc ion bridge chelated by histidine side-chains and connected to the cytoplasmic ends of Helix 3 and 6 is observed to prevent receptor activation. This perhaps indicates that separation of these cytoplasmic ends would contribute to rhodopsin activation[14].

The structure of rhodopsin may provide stability to the important Schiff base linkage with the retinal by affecting its hydrolysis, limiting its interactions with solvent, and inhibiting its release when hydrolyzed, thus encouraging rebinding of the Schiff base linkage[15].

Retinal Chromophore of Rhodospin

Rhodopsin is bound covalently to the 11-cis retinal, the chromophore or "ligand," and this retinal is found in deeply in the core of the helices, in a hydrophobic site, parallel to the lipid bilayer[16]. The retinal is attached in the active site of rhodopsin through a protonated Schiff base bond to the ε-amino group of Lysine 296 residue on the C-terminal Helix 7, with this linkage creating a positive charge on the chromophore [17]. As this ligand is bound in the 12-s-trans conformation, there arises the non-bonding interactions between the C-13 methyl group and C-10 hydrogen that contribute to non-planarity. This leads to the ability of the chromophore polyene tail to undergo fast photoisomerization around the C-11=C-12 double bond during light-induced activation[18]. Somewhat enclosing this chromophore is a retinal binding pocket partially formed by the N-terminal domain overlaying the extracellular turns including Extracellular Helix 2, which folds into the molecular center[19].

Function

Light-Induced Visual Signal Transduction

In the active site of rhodopsin, there is a protonated 11-cis retinal (retinylidene) Schiff base which functions as an inverse agonist. In photoisomeration, the primary step in rhodopsin photoactivation, 11-cis retinylidene ligand is switched into an all-trans retinal configuration. In this <200 fs process, the protein-binding pocket accommodates the 11-cis conformation of the chromophore and is preserved, restraining the relaxation of the chromophore. The strained chromophore relaxation of conformational energy transforms the protein from the inactive to active form[20].

Rhodopsin is converted into the active signalling state of metarhodopsin MII. Rhodopsin kinase phosphorylates rhodopsin and arrestin binds to rhodopsin, preventing further signal transduction from MII. The all-trans¬-retinal Schiff base linkage of MII may be hydrolyzed and cleaved, releasing free retinal and leaving the apoprotein, opsin. Free retinal can be enzymatically converted into 11-cis-retinal form, then rebinds to opsin to regenerate rhodopsin[21] When metarhodopsin II is formed, Schiff base deprotonates and proton is transferred to Glu113, releases stored light energy and activates the protein. The retinal protonated Schiff base and Glu113 efficiently locks the receptor in an inactive state in the dark.

Rhodopsin can convert light energy into a chemical signal immensely efficiently.[22]

Signal transduced is a “ligand” consisting of a photon interacting with a covalently bound retinal, changing the conformation of the retinal and opsin (protein moiety). This generates a signalling cascade where a large number of transducin molecules, the trimeric G protein in the cytoplasm, are activated through trigger of GDP-GTP nucleotide exchange in the α subunit. This results in the hyperpolarisation of the rod photoreceptor cells and synaptic signalling to adjacent rod bipolar cells; each activated transducin activates cGMP-specific phosphodiesterase, which results in the hydrolysis of cGMP molecules and the closing of the cGMP-gated cation channels in the plasma membrane of the outer segment. The cell hyperpolarises due to the decrease in the influx of sodium and calcium ions, results in the decrease in the release of glutamate into the synaptic cleft. ....For the post-synaptic state, the reduced glutamate levels inhibit G protein-mediated signalling by mGluR6 receptors resulting in the opening of the cation channels, depolarisation, and transmittance of the signals from the retina through nerve impulses to the visual system. 11-cis-retinal seems to act as an inverse agonist, repressing opsin activity, stabilizing the structural inactive conformation. When activated by light, a photochemical reaction occurs in picoseconds where the retinal is converted into an all-trans form. Following that is a slower thermal relaxation process involving retinal and opsin undergoing conformational changes to result in fully active metarhodopsin II. [23].

It phosphorylates both metarhodopsin I and II and cone opsins. The majority of the phosphorylation sites were in the cytoplasmic C-terminal region of rhodopsin with seven hydroxy-amino acids. Most favoured were Ser 338, Ser 343, Ser 334, Thr 335 and Thr 336. Protein phosphatise 2A recognizes and dephosphorylates rhodopsin.[24].


The 11-cis-retinal is permanently bound to the receptor as a prosthetic group and inactivates it. The retinal plays a chromophore in the initial photochemistry state and as an agonist after the relaxation state in producing the active state of the receptor. To desensitize signalling to a G protein, phosphorylating the activated receptor is one method. The major phosphorylation sites of rhodopsin are Ser 338, Ser 343 and Ser 334. These residues form an arrangement that does not appear to be exposed to the solvent. Interactions with the C-terminal tail and a portion of the C-III loop appear to be broken for the phosphorylation of the hydroxyl groups. 1) Light induced isomerization of the retinal from cis to trans. Fits well in retinal pocket. 2) thermal relaxation of the retinal-protein complex 3) Late equilibria affected by rhodopsin and G protein interaction. The meta states: Metarhodopsin I forms to Metarhodopsin II which is the intermediate signalling state where interaction occurs with the G protein. This is accompanied by the breakage of the salt bridge between Glu 113 and the protonated Schiff base between Lys 296 and retinal, and movement in the helices. Occurs in milliseconds. The Schiff base proton is transferred to the Glu 113 counterion., thus destabilizing the ground state. The cytoplasm also uptakes protons in the formation of MII. Is speculated to be dependent on the protonation of Glu134, a highly conserved residue, which destabilizes the constraint on the Arg 135 it is salt-bridged with. In forming the signalling state, there is positive enthalpy with the formation of MII. Formation of the active state, linked with the increase in entropy, is suggested to release the constrains in the helices and expose the cytoplasmic binding sites. The 9-methyl group of retinal is suggested to provide a scaffold for proton transfers essential for the formation of the active state[25].

Like most GCPRs though, the activated rhodopsin catalyzes uptake of GTP by heterotrimeric G protein, in this case transducin, which interacts with cytoplasmic loops of the receptor. Dissociation of the GTP-bound α-subunit from the βγ heterodimer enables triggering of the rod cell’s response to light. Upon activation, movement and slight adjustment of helices are observed, with the inner faces of Helix II, III, VI and VII more exposed[26].

Different states including short-lived, photo- batho- and lumi-rhodopsin, and longer-lived meta-rhodopsins give information of the structural status of the molecule during activation. Upon activation, Meta II state formation is accompanied by deprotonation of the Schii base, breakage of the salt bridge with couterion residue Glu 113. Upon activation, helices three and six move outwards, binding site for transducin is accessible as opening between cytoplasmic loops. Deactivation occurs when rhodopsin kinase phosphorylates rhodopsin’s C-terminal region, and arrestin binds, competing with transducin. Proceeds by decay of Meta II to opsin. For activation of rhodopsin, rhodopsin has to be dephosphorylated, and have the 11-cis retinal form again[27].


Opsin

References

  • Okada T, Sugihara M, Bondar AN, Elstner M, Entel P, Buss V. The retinal conformation and its environment in rhodopsin in light of a new 2.2 A crystal structure. J Mol Biol. 2004 Sep 10;342(2):571-83. PMID:15327956 doi:10.1016/j.jmb.2004.07.044
  1. ↑ Article 1
  2. ↑ Article 12
  3. ↑ Article 20
  4. ↑ Article 20
  5. ↑ Article 20
  6. ↑ Article 12
  7. ↑ Article 4
  8. ↑ Article 9
  9. ↑ Article 9
  10. ↑ Article 12
  11. ↑ Article 9
  12. ↑ Article 12
  13. ↑ Article 20
  14. ↑ Article 10
  15. ↑ Article 3
  16. ↑ Article 19
  17. ↑ Article 4
  18. ↑ Article 2
  19. ↑ Article 6
  20. ↑ Article 2
  21. ↑ Article 3
  22. ↑ Article 4
  23. ↑ Article 6
  24. ↑ Article 7
  25. ↑ Article 9
  26. ↑ Article 10
  27. ↑ Article 19
Please do NOT make changes to this Sandbox until after April 23, 2010. Sandboxes 151-200 are reserved until then for use by the Chemistry 307 class at UNBC taught by Prof. Andrea Gorrell.
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