Rhodopsin GPCR

Katharina Devitofranceschi '14 Noah Winters '15


I. Introduction

Vision is a crucial element to almost all facets of mammalian life. For most animals it is required to find food, interact with conspecifics and locate home. At the most fundamental level, rod and cone cells in mammalian retina confer the ability of vision. The process begins with the absorption of light via rod cells and the G-protein coupled receptor (GPCR), rhodopsin. Detection of the photon occurs through an extremely fast, highly selective and efficient reaction mediated by a conformational change in 11-cis-retinal. This opsin/photon reaction then cues a cascade of signals that excite neurons involved in vision and allow for the perception of an image. The rhodopsin GPCR represents a paradigm for the structural functions of these receptor types . Out of the 2-3% of mammalian genes that code for GPCRs, approximately 90% of those GPCRs belong to the rhodopsin family. As such, an understanding for rhodopsin's structural function is paramount.  

II. General Structure

    All GPCRs contain highly conserved 7-transmembrane helices.

  In rhodopsin, helices I, IV, VI, and VII are all kinked due to Pro resides, though only significantly at helices IV and VI.  Helix VII possess a structural irregularity resulting from the binding of Lys296 to the 11-cis-retinal chromophore. Rhodopsin contains a cytoplasmic terminal region, consisting of helix II ( Pro71 and Leu72), C-II (Phe148), helix V (Leu226, Val230), and helix VI (Val250, Met253). Together, this region forms the binding and activation site for a G protein.

Rhodopsin contains an extracellular domain comprised of the NH2 terminal and interhelical loops I, II, and III. The NH2-terminal tail is composed of 5 strands, the first two being antiparallel beta sheets (Gly3-Pro12) which run almost parallel to the phospholipid membrane. The other three strands run from Phe13 to Pro34.

The E-I and E-III loops run along the periphery of the molecule, while the middle of the E-II loop penetrates deep inside the GPCR with two antiparallel beta sheets. The uppermost sheet forms part of the chromophore-binding pocket  

III. 11-cis-Retinal Binding

As aforementioned, the 11-cis-Retinal chromophore is attached to Lys296. The residues interact via a Schiff base linkage, as indicated by the merging of the densities of chromophore's polyene chain and the side chain of Lys296. Retinal is located closer to the extracellular region of the lipid bilayer, rather than the interdiscal region. The portion of the binding pocket that surrounds the beta-ionone ring of retinal contains residues that are close to the cytoplasmic side of the membrane. These residues include Glu122, Phe261,and Trp265, as well as the residues Met207, His211, Phe212, Tyr268, Ala269 from helix VI. A kink introduced by Pro267 causes these residues to cover the beta-ionone ring within the pocket, binding non-specifically. Binding of the polyene chain within the binding pocket is also done through non-specific interactions with the residues Glu113, Gly114, Ala117, Thr118, Gly120, and Gly121 , Cys167, Tyr43, Met44, Leu47, and the beta sheet from EII. The unique orientation of Lys296 is directed by the hydrophobic residues Met44 and Leu47, and the peptide bond between Phe293 and Phe294. The entire area is stabilized by the two phenyl rings interacting with adjacent helices II and VI. Counterion formation and subsequent Schiff linkage stabilization are facilitated by Glu113 and Thr94. There is a distance of 3.3 and 3.5 between the carboxylate oxygen atoms of Glu113 and Thr94, and the nitrogen atom of the Schiff base.


IV. Photoactivation of Rhodopsin

When light energy in the form of photons hits the 11-cis-retinal chromophore, the molecule isomerizes into its all-trans conformation. This isomerization results in several changes in binding affinity within the receptor. First, the beta-ionone ring moves towards helix III, and is accompanied by displacement of the C9 and C13 methyl groups of retinal. Movement of the methyl regions results in a transformation of the salt-bridge between Glu113 and the Schiff base. This action results in neutralization of the previously charged species and displacement of helix III. Movement of helix III disrupts the binding between Glu122 and His211, as well as the C13 methyl of retinal and Trp265. Furthermore, photoactivation and trans-isomerization leads to the splitting of interhelical and hydrophobic constraints, mediated by Ala299, Asn302, and Tyr306, and Phe294, respectively. As a result, the receptor   undergoes conformational rearrangement which results in subsequent activation of a cytoplasmic G protein.

After the chromophore is converted to its all-trans conformation, the molecule is released from the receptor into the cytoplasm. The four cytoplasmic-facing residues Lys67, Lys66 , Arg69, and His65 mediate this release.  

V.Rhodopsin and Other GPCRs

Rhodopsin is considered to be the paradigm of structural GPCR studies. But how similar is it really to other GPCRs?

Interestingly, there is a great deal of extracellular structural divergence. The N terminus of rhodopsin along with the extracellular loop 2 (ECL2) forms a four-stranded beta-sheet. This beta-sheet additionally interacts with the ECL1 and ECL3. These structures serve to occlude the binding site from other ligands.

In comparison, the beta-2-adrenergic receptor   (which catalyzes the crucial epinehrine signaling cascade) is structurally very open, and is able to bind several different types of ligands. The primary feature of the beta-2AR is a short helical segment within the ECL2. This helix is supported by a few di-sulfide interactions and contact with ECL1.

The transmembrane region is the most conserved sequence between GPCRs. They all share a common structural core of 97 residues. As a result, the helical bundle orientation is similar across all 4 crystallized GPCRs to date.

It stands to reason that the ligand-binding pocket is what varies most between GPCRs. Surprisingly, though, the beta-2-adrenergic receptor has a pocket that structurally resembles that of rhodopsin. The position of the pocket is fairly similar and in both cases the ligand binding extends from TM VII. In beta-2AR the ligand engages in a strong polar interaction while in rhodopsin this interaction is a full-fledged covalent bond.

Knowing the structural differences between GPCRs is crucial to understanding the most ubiquitous receptor type in our body, and is of particular significance as a pharmacological target for drug therapies.

VI. References

Hanson, M. A., & Stevens, R. C. (2009). "Discovery of new GPCR biology: one receptor structure at a time." Structure,17(1), 8-14.

Jung Hee Park, Patrick Scheerer, Klaus Peter Hofmann, Hui-Woog Choe & Oliver Peter Ernst (2008)."Crystal structure of the ligand-free G-protein-coupled receptor opsin" Nature, 454 183-187

Palczewski, K., Kumasaka, T., Hori, T., Behnke, C. A., Motoshima, H., Fox, B. A., & Miyano, M. (2000)."Crystal structure of rhodopsin: AG protein-coupled receptor." Science Signaling, 289(5480), 739.

Okada, T., Sugihara, M., Bondar, A. N., Elstner, M., Entel, P., & Buss, V. (2004). "The retinal conformation and its environment in rhodopsin in light of a new 2.2 crystal structure." Journal of Molecular Biology, 342(2), 571-583.

Teller, D.C.,Okada, T., Stenkamp, R.E., "Advances in Determination of a High-Resolution Three-Dimensional Structure of Rhodopsin, a Model of G-Protein-Coupled Receptors (GPCRs)" Biochemistry, 40(26):7761-7772

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