Goldberg A F, Molday R S
Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, Canada.
Biochemistry. 1996 May 14;35(19):6144-9. doi: 10.1021/bi960259n.
Peripherin/rds and rom-1 are homologous integral membrane protein subunits found as an oligomeric complex at the rim regions of rod and cone photoreceptor outer segment disks. These proteins are essential for the morphogenesis of normal outer segments and have been linked to a variety of human retinal degenerative diseases. Previous studies have suggested that disulfide-linked homodimers of peripherin/rds and rom-1 can associate noncovalently to form higher order structures. We have characterized the hydrodynamic properties of Triton X-100 solubilized peripherin/rds-rom-1 complexes from bovine ROS membranes by gel exclusion chromatography on Sepharose C1-6B and velocity sedimentation through H2O- and D2O-based sucrose gradients. A single hydrodynamic species is observed which has a Stokes radius of 6.2 nm, a sedimentation coefficient (S20,w) of 5.8 S, and a partial specific volume of 0.83 mL/g. From these data the molecular mass of the detergent-peripherin/rds-rom-1 complex is calculated to be 240 kDa. The protein component of this complex is estimated to be 135 kDa, providing direct evidence that the solubilized peripherin/rds-rom-1 complex is a tetramer. The abundance of this complex as measured by competitive ELISA and immunoaffinity purification is approximately 4% of total bovine ROS membrane protein and indicates that peripherin/rds-rom-1 tetramers are present at a relatively high average surface density (ca. 4100/ microns m2) at the rim surfaces of rod outer segment disks.
外周蛋白/视网膜变性慢病毒（Peripherin/rds）和视网膜外节膜蛋白1（rom-1）是同源整合膜蛋白亚基，以寡聚复合物的形式存在于视杆和视锥光感受器外节盘膜的边缘区域。这些蛋白质对于正常外节的形态发生至关重要，并与多种人类视网膜退行性疾病有关。先前的研究表明，外周蛋白/rds和rom-1的二硫键连接的同型二聚体可以非共价结合形成更高阶的结构。我们通过在琼脂糖C1-6B上进行凝胶排阻色谱以及通过基于H2O和D2O的蔗糖梯度进行速度沉降，表征了来自牛视网膜外节膜（ROS）膜的Triton X-100溶解的外周蛋白/rds-rom-1复合物的流体动力学性质。观察到单一的流体动力学物种，其斯托克斯半径为6.2 nm，沉降系数（S20,w）为5.8 S，偏比容为0.83 mL/g。根据这些数据，去污剂-外周蛋白/rds-rom-1复合物的分子量计算为240 kDa。该复合物的蛋白质成分估计为135 kDa，这直接证明溶解的外周蛋白/rds-rom-1复合物是四聚体。通过竞争性酶联免疫吸附测定（ELISA）和免疫亲和纯化测量，该复合物的丰度约为牛ROS膜总蛋白的4%，表明外周蛋白/rds-rom-1四聚体以相对较高的平均表面密度（约4100/μm2）存在于视杆外节盘膜的边缘表面。
