ReviewCrystal structure of rhodopsin: implications for vision and beyond
Introduction
G-protein-coupled receptors (GPCRs) are involved in a number of physiological processes, such as sensory transduction, mediation of hormonal activity and cell-to-cell communication. In general, GPCRs are activated as a result of the binding of extracellular, receptor-specific ligands to their extracellular or transmembrane domains. Conformational changes of the receptor induced by ligand binding are relayed to the cytoplasmic surface, allowing productive coupling of the receptor with a heterotrimeric G protein. In this context, visual photoreceptor proteins, called visual pigments, are unique in that their intrinsic ligand, 11-cis-retinal, is coupled via a protonated Schiff base (PSB) to a sidechain of a transmembrane lysine residue (Lys296). The 11-cis-retinylidene moiety acts as an inverse agonist, suppressing the activity of the receptor to an undetectable level. This feature is especially important for the function of rod cells, which contain approximately 3 mM of the visual photoreceptor protein rhodopsin (108 molecules) and are specialized to detect light at very low rates of photon absorption. Any spontaneous activation of rhodopsin present at such a high concentration would trigger phototransduction, lowering the overall light sensitivity 1., 2., 3.. The ground state of rhodopsin, with its intrinsic ligand and its rigid extracellular structure, appears to be specifically suited to maintaining the receptor in the inactive state.
From a structural point of view, GPCRs are defined by seven transmembrane α helices (7TMH) (I–VII) connected by three extracellular (E-I–E-III) and three intracellular (C-I–C-III) loops. The largest subfamily of GPCRs is called the rhodopsin family because its members share many residues and motifs with rhodopsin in their transmembrane regions. More importantly, a common activation mechanism that involves rearrangement of the 7TMH 4., 5. is expected for at least some of the members of the rhodopsin family [6]. In the case of bovine rhodopsin, photoactivation causes such significant structural changes that the three-dimensional crystal lattice is no longer maintained [7•] (Fig. 1).
Historically, based on electron microscopy studies, the 7TMH bundle structure was first proposed for bacteriorhodopsin (bR) [8], which is functionally distinct from 7TMH rhodopsin. Consequently, rhodopsin, bR and other bacterial proteins are often grouped as retinal proteins (or, more correctly, retinylidene-binding proteins), even though the chromophore of the bacterial proteins is all-trans-retinal 9., 10., 11..
Significant progress in structural studies of bacterial retinal proteins has been made during the past few years (reviewed 9., 10., 11.) 12., 13., 14., 15., 16., 17.. These results have yielded detailed structural information for ion pumps, but have a limited value for understanding the GPCR structure/function relationship, because there is no evolutionary relationship (sequence similarity, gene organization, etc.) between bacterial retinal proteins and rhodopsin or any of the GPCRs. Moreover, the protein-specific photoisomerization reactions of the retinal chromophore in rhodopsin and bR lead to unique conformational changes for both receptors. Additionally, there are other proteins that are light sensitive. Most noteworthy, photoactive yellow protein (PYP), a soluble nonretinal protein, contains a 4-hydroxycinnamic acid chromophore, which photoisomerizes around the 7,8-vinyl bond during the initial step in its conformational transition to the signaling state 18., 19..
X-ray crystallography [20••] and further refinement of an initial model of rhodopsin [21••] provide the first structural information about the organization of the polypeptide chain and the post-translational moieties in the ground state of rhodopsin. These studies are especially valuable both for understanding the molecular constraints of rhodopsin that prevent spontaneous activation achieving the extremely low noise in rod cells and for identifying residues that directly interact with the chromophore. The intimate hydrophobic contacts between residues of the protein (called opsin) and the chromophore are vital to the spectral tuning of other visual pigments that allow color discrimination (color vision). In this review, we will focus on how the rhodopsin structural information can be applied to a large number of GPCRs; we propose that there is a correspondence of the basic architecture and a common activation mechanism. We will also discuss how the primary photoactivation events of rhodopsin may be similar to events during photoactivation of non-GPCRs, bRs and PYP.
Section snippets
Crystal structure of bovine rhodopsin
The molecular architecture of rhodopsin is purposely designed to support the function of this receptor. Rhodopsin has minimal activity in the dark and efficiently captures light energy sufficient to overcome thermodynamically unfavorable changes in the protein, including the rearrangement of the cytoplasmic surface, thus allowing interaction with G proteins. Rhodopsin contains its chromophore ligand covalently linked to helix VII. This arrangement stabilizes the helical bundle in the off
The chromophore trigger in rhodopsin
Switching from the 11-cis- to the all-trans-retinylidene conformation by photon absorption, within milliseconds, forces the protein to assume a distinct active conformation that is competent to catalyze the GDP/GTP exchange reaction on the α subunit of the G protein transducin (Gt), which is already coupled to rhodopsin. The importance of the chromophore in this process has been emphasized in recent studies [24•]. Initial changes to the chromophore–protein interaction most probably involve the
Activation mechanism for G-protein-coupled receptors
The changes to the chromophore–protein interaction described above lead to the rearrangement of the cytoplasmic side of the 7TMH. The major intramolecular constraints were found to be hydrophobic interactions surrounding an E(D)RY(W) (3.49–3.51) motif, which is highly conserved among receptors of the rhodopsin family of GPCRs (Fig. 4). A number of mutational studies have shown the direct involvement of this motif in the activation process of rhodopsin [40] and other ligand-binding receptors [6].
Conclusions
The crystal structure of bovine rhodopsin provides a new point from which a number of previous structural/functional studies on a variety of GPCRs can be re-examined and new studies conceptualized. More importantly, it will help to design future experimental and theoretical approaches to explore further details about these receptors. Moreover, future crystallographic studies on the photoactivation mechanism of rhodopsin could contribute to increasing our knowledge not only of the
Update
Recent work [51] has suggested that the native chromophore cis-trans isomerization is initially required to store light energy in the opsin–chromophore complex. The energy is subsequently utilized for repositioning the β-ionone ring, helix movement and receptor activation. Furthermore, illumination of the active forms of rhodopsin generates a product that is in an inactive conformation, but has an all-trans-retinal chromophore in the binding pocket [52]. This suggests that, to regenerate
Acknowledgements
We would like to thank PA Hargrave, HE Hamm and Naomi Wilson for their comments on this manuscript. This research was supported by National Institutes of Health grant EY-09339, awards from Research to Prevent Blindness, Inc. (RPB) to the Department of Ophthalmology at the University of Washington, the Foundation Fighting Blindness, the Ruth and Milton Steinbach Fund, and the EK Bishop Foundation. KP is an RPB Senior Investigator.
References and recommended reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
• of special interest
•• of outstanding interest
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