Department of Pharmacology, Vanderbilt University, Nashville, Tennessee, United States.
Human Genetics Center, School of Public Health, The University of Texas Health Science Center, Houston, Texas, United States.
Invest Ophthalmol Vis Sci. 2018 Jan 1;59(1):13-20. doi: 10.1167/iovs.17-22180.
The purpose of this study was to identify the molecular defect in the disease-causing human arrestin-1 C147F mutant.
The binding of wild-type (WT) human arrestin-1 and several mutants with substitutions in position 147 (including C147F, which causes dominant retinitis pigmentosa in humans) to phosphorylated and unphosphorylated light-activated rhodopsin was determined. Thermal stability of WT and mutant human arrestin-1, as well as unfolded protein response in 661W cells, were also evaluated.
WT human arrestin-1 was selective for phosphorylated light-activated rhodopsin. Substitutions of Cys-147 with smaller side chain residues, Ala or Val, did not substantially affect binding selectivity, whereas residues with bulky side chains in the position 147 (Ile, Leu, and disease-causing Phe) greatly increased the binding to unphosphorylated rhodopsin. Functional survival of mutant proteins with bulky substitutions at physiological and elevated temperature was also compromised. C147F mutant induced unfolded protein response in cultured cells.
Bulky Phe substitution of Cys-147 in human arrestin-1 likely causes rod degeneration due to reduced stability of the protein, which induces unfolded protein response in expressing cells.
本研究旨在确定导致疾病的人 arrestin-1 C147F 突变体的分子缺陷。
测定野生型（WT）人 arrestin-1 和几个位置 147 取代突变体（包括导致人类显性视网膜炎色素变性的 C147F）与磷酸化和非磷酸化光激活视紫红质的结合。还评估了 WT 和突变型人 arrestin-1 的热稳定性以及 661W 细胞中的未折叠蛋白反应。
WT 人 arrestin-1 对磷酸化光激活视紫红质具有选择性。用较小侧链残基丙氨酸或缬氨酸取代 Cys-147 不会显著影响结合选择性，而在位置 147 处具有较大侧链的残基（异亮氨酸、亮氨酸和致病的苯丙氨酸）则大大增加了对非磷酸化视紫红质的结合。在生理和升高温度下，具有大取代的突变蛋白的功能存活也受到影响。C147F 突变体在培养细胞中诱导未折叠蛋白反应。
人 arrestin-1 中 Cys-147 的大苯丙氨酸取代可能由于蛋白质稳定性降低而导致杆状细胞变性，从而在表达细胞中诱导未折叠蛋白反应。
