title: Molecular mechanisms of disease for mutations at Gly-90 in rhodopsin.
abstract: Two different mutations at Gly-90 in the second transmembrane helix of the photoreceptor protein rhodopsin have been proposed to lead to different phenotypes. G90D has been classically associated with congenital night blindness, whereas the newly reported G90V substitution was linked to a retinitis pigmentosa phenotype. Here, we used Val/Asp replacements of the native Gly at position 90 to unravel the structure/function divergences caused by these mutations and the potential molecular mechanisms of inherited retinal disease. The G90V and G90D mutants have a similar conformation around the Schiff base linkage region in the dark state and same regeneration kinetics with 11-cis-retinal, but G90V has dramatically reduced thermal stability when compared with the G90D mutant rhodopsin. The G90V mutant also shows, like G90D, an altered photobleaching pattern and capacity to activate Gt in the opsin state. Furthermore, the regeneration of the G90V mutant with 9-cis-retinal was improved, achieving the same A(280)/A(500) as wild type isorhodopsin. Hydroxylamine resistance was also recovered, indicating a compact structure around the Schiff base linkage, and the thermal stability was substantially improved when compared with the 11-cis-regenerated mutant. These results support the role of thermal instability and/or abnormal photoproduct formation in eliciting a retinitis pigmentosa phenotype. The improved stability and more compact structure of the G90V mutant when it was regenerated with 9-cis-retinal brings about the possibility that this isomer or other modified retinoid analogues might be used in potential treatment strategies for mutants showing the same structural features.
authors.affiliation: Centre de Biotecnologia Molecular, Departament d'Enginyeria Química, Universitat Politècnica de Catalunya, 08222 Terrassa, Spain.
authors.forename: Darwin
authors.initials: D
authors.lastname: Toledo
authors.forename: Eva
authors.initials: E
authors.lastname: Ramon
authors.forename: Mònica
authors.initials: M
authors.lastname: Aguilà
authors.forename: Arnau
authors.initials: A
authors.lastname: Cordomí
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Pérez
authors.forename: Hugo F
authors.initials: HF
authors.lastname: Mendes
authors.forename: Michael E
authors.initials: ME
authors.lastname: Cheetham
authors.forename: Pere
authors.initials: P
authors.lastname: Garriga
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002417:Cattle; D045744:Cell Line, Tumor; D004224:Diterpenes; D015785:Eye Diseases, Hereditary; D040181:Genetic Diseases, X-Linked; D006801:Humans; D020125:Mutation, Missense; D009216:Myopia / Q000235:genetics / Q000378:metabolism*; D009755:Night Blindness / Q000235:genetics / Q000378:metabolism*; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D012172:Retinaldehyde / Q000235:genetics / Q000378:metabolism; D012174:Retinitis Pigmentosa / Q000235:genetics / Q000378:metabolism*; D012243:Rhodopsin / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D004224:Diterpenes; C031390:9-cis-retinal; D012243:Rhodopsin; D012172:Retinaldehyde