title: Improved conformational stability of the visual G protein-coupled receptor rhodopsin by specific interaction with docosahexaenoic acid phospholipid.
abstract: Rhodopsin is the photoreceptor located in the rod cells of the retina. It has seven transmembrane helices and is a prototypic member of the G protein-coupled receptor superfamily. The structures and functions of these receptors are clearly affected by the lipid composition of the cell membrane, and their study in a purified recombinant form is usually performed in detergent solution. There is a need to study these receptors in a physiologically relevant environment because the lipid environment is known to have an important effect on their function. In this work, rhodopsin reconstituted in docosahexaenoic acid (DHA) liposomes is shown to have more thermal stability than when it is solubilised with the neutral detergent dodecyl maltoside. Moreover, the specific interaction between rhodopsin and DHA was followed by means of Langmuir experiments with insertion of rhodopsin into lipid monolayers; this showed high affinity for the lipid-receptor interaction. Furthermore, fluorescence spectroscopy measurements indicate that the conformation of opsin obtained after photobleaching is preserved in DHA-containing liposomes, thereby allowing retinal to re-enter the binding pocket even long after bleaching. Overall, our results demonstrate that liposomes of this specific lipid provide a more stable environment for ground-state inactive rhodopsin in the dark, than dodecyl maltoside detergent, and that this lipid can also preserve the native correctly folded ligand-free opsin conformation obtained after illumination. This strategy will be used in further studies on mutations of rhodopsin associated with congenital retinopathies.
authors: Chemical Engineering Department, Centre de Biotecnologia Molecular, Universitat Politècnica de Catalunya, Edifici Gaia, Rambla de Sant Nebridi s/n, 08222 Terrassa, Catalonia, Spain.
authors: Maria Jesús
authors: MJ
authors: Sánchez-Martín
authors: Eva
authors: E
authors: Ramon
authors: Juan
authors: J
authors: Torrent-Burgués
authors: Pere
authors: P
authors: Garriga
mesh_terms: D004281:Docosahexaenoic Acids / Q000737:chemistry*; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism*; D038761:Photobleaching; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012243:Rhodopsin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature
chemical_list: D008081:Liposomes; D043562:Receptors, G-Protein-Coupled; D011994:Recombinant Proteins; D004281:Docosahexaenoic Acids; D012243:Rhodopsin