ジャーナル: Proc Natl Acad Sci U S A / 年: 2016 タイトル: Paired octamer rings of retinoschisin suggest a junctional model for cell-cell adhesion in the retina. 著者: Gökhan Tolun / Camasamudram Vijayasarathy / Rick Huang / Yong Zeng / Yan Li / Alasdair C Steven / Paul A Sieving / J Bernard Heymann / 要旨: Retinoschisin (RS1) is involved in cell-cell junctions in the retina, but is unique among known cell-adhesion proteins in that it is a soluble secreted protein. Loss-of-function mutations in RS1 lead ...Retinoschisin (RS1) is involved in cell-cell junctions in the retina, but is unique among known cell-adhesion proteins in that it is a soluble secreted protein. Loss-of-function mutations in RS1 lead to early vision impairment in young males, called X-linked retinoschisis. The disease is characterized by separation of inner retinal layers and disruption of synaptic signaling. Using cryo-electron microscopy, we report the structure at 4.1 Å, revealing double octamer rings not observed before. Each subunit is composed of a discoidin domain and a small N-terminal (RS1) domain. The RS1 domains occupy the centers of the rings, but are not required for ring formation and are less clearly defined, suggesting mobility. We determined the structure of the discoidin rings, consistent with known intramolecular and intermolecular disulfides. The interfaces internal to and between rings feature residues implicated in X-linked retinoschisis, indicating the importance of correct assembly. Based on this structure, we propose that RS1 couples neighboring membranes together through octamer-octamer contacts, perhaps modulated by interactions with other membrane components.
全体 : Mature retinoschisin with a C-terminal hexahistidine tag
全体
名称: Mature retinoschisin with a C-terminal hexahistidine tag
要素
試料: Mature retinoschisin with a C-terminal hexahistidine tag
タンパク質・ペプチド: Retinoschisin 1
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超分子 #1000: Mature retinoschisin with a C-terminal hexahistidine tag
超分子
名称: Mature retinoschisin with a C-terminal hexahistidine tag タイプ: sample / ID: 1000 / 集合状態: 16-mer / Number unique components: 1
分子量
実験値: 400 KDa / 理論値: 382 KDa / 手法: Blue native gel electrophoresis
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分子 #1: Retinoschisin 1
分子
名称: Retinoschisin 1 / タイプ: protein_or_peptide / ID: 1 / Name.synonym: RS1 詳細: The mature RS1 protein (no signal sequence) with a C-terminal six-histidine tag コピー数: 16 / 集合状態: 16-mer / 組換発現: Yes
由来(天然)
生物種: Homo sapiens (ヒト) / 別称: Human / 組織: Retina / 細胞: Photoreceptors and bipolar cells / Organelle: Extracellular space / 細胞中の位置: Forming a junction between cells
詳細: C-flat holey carbon grids (2.0 um holes, 2.0 um spaces)
凍結
凍結剤: ETHANE / チャンバー内湿度: 90 % / 装置: LEICA EM GP / 詳細: Plunged at room temperature. / 手法: Blotted from the copper side.
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電子顕微鏡法
顕微鏡
FEI POLARA 300
日付
2014年6月11日
撮影
カテゴリ: CCD / フィルム・検出器のモデル: GATAN K2 (4k x 4k) / デジタル化 - サンプリング間隔: 5 µm / 実像数: 19 / 平均電子線量: 30 e/Å2 詳細: Each image is the average of 10 frames taken over 3 seconds.
電子線
加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN
電子光学系
照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / Cs: 2.0 mm / 倍率(公称値): 39000
試料ステージ
試料ホルダーモデル: OTHER
実験機器
モデル: Tecnai Polara / 画像提供: FEI Company
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画像解析
詳細
Particles were picked semi-automatically (EMAN2).
CTF補正
詳細: Each particle
最終 再構成
解像度のタイプ: BY AUTHOR / 解像度: 4.1 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: Relion, EMAN2, Bsoft / 使用した粒子像数: 9096