Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, Max-von-Laue-Strasse 7, 60438, Frankfurt, Germany.
Institute for Organic Chemistry and Chemical Biology, Goethe University Frankfurt, Max-von-Laue-Strasse 7, 60438, Frankfurt, Germany.
Angew Chem Int Ed Engl. 2020 Sep 1;59(36):15656-15664. doi: 10.1002/anie.202003671. Epub 2020 Aug 13.
The RHO gene encodes the G-protein-coupled receptor (GPCR) rhodopsin. Numerous mutations associated with impaired visual cycle have been reported; the G90D mutation leads to a constitutively active mutant form of rhodopsin that causes CSNB disease. We report on the structural investigation of the retinal configuration and conformation in the binding pocket in the dark and light-activated state by solution and MAS-NMR spectroscopy. We found two long-lived dark states for the G90D mutant with the 11-cis retinal bound as Schiff base in both populations. The second minor population in the dark state is attributed to a slight shift in conformation of the covalently bound 11-cis retinal caused by the mutation-induced distortion on the salt bridge formation in the binding pocket. Time-resolved UV/Vis spectroscopy was used to monitor the functional dynamics of the G90D mutant rhodopsin for all relevant time scales of the photocycle. The G90D mutant retains its conformational heterogeneity during the photocycle.
RHO 基因编码 G 蛋白偶联受体（GPCR）视紫红质。已经报道了许多与视觉周期受损相关的突变；G90D 突变导致视紫红质的组成型激活突变形式，从而导致 CSNB 疾病。我们通过溶液和 MAS-NMR 光谱报告了在黑暗和光激活状态下结合口袋中视网膜构型和构象的结构研究。我们发现 G90D 突变体与 11-顺式视黄醛结合作为希夫碱在两种群体中都有两个长寿命的暗状态。在黑暗状态下的第二个次要群体归因于结合口袋中盐桥形成引起的突变诱导的扭曲导致共价结合的 11-顺式视黄醛构象的轻微变化。时间分辨的紫外/可见光谱用于监测 G90D 突变体视紫红质在光循环的所有相关时间尺度上的功能动力学。G90D 突变体在光循环过程中保持其构象异质性。
