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基本情報
登録情報 | データベース: PDB / ID: 4v4n | |||||||||
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タイトル | Structure of the Methanococcus jannaschii ribosome-SecYEBeta channel complex | |||||||||
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![]() | RIBOSOME/PROTEIN TRANSPORT / archaea / archaeal / ribosomal / 50S / protein synthesis / RNA / large subunit / co-translational translocation / protein conducting channel / RIBOSOME-PROTEIN TRANSPORT complex | |||||||||
機能・相同性 | ![]() intracellular protein transmembrane transport / signal sequence binding / SRP-dependent cotranslational protein targeting to membrane, translocation / protein transmembrane transporter activity / protein secretion / protein targeting / protein transport / plasma membrane 類似検索 - 分子機能 | |||||||||
生物種 | ![]() ![]() | |||||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 9 Å | |||||||||
![]() | Menetret, J.F. / Park, E. / Gumbart, J.C. / Ludtke, S.J. / Li, W. / Whynot, A. / Rapoport, T.A. / Akey, C.W. | |||||||||
![]() | ![]() タイトル: Structure of the SecY channel during initiation of protein translocation. 著者: Eunyong Park / Jean-François Ménétret / James C Gumbart / Steven J Ludtke / Weikai Li / Andrew Whynot / Tom A Rapoport / Christopher W Akey / ![]() 要旨: Many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic SecY or eukaryotic Sec61 complexes, and are translocated across the membrane during ...Many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic SecY or eukaryotic Sec61 complexes, and are translocated across the membrane during their synthesis. Crystal structures of the inactive channel show that the SecY subunit of the heterotrimeric complex consists of two halves that form an hourglass-shaped pore with a constriction in the middle of the membrane and a lateral gate that faces the lipid phase. The closed channel has an empty cytoplasmic funnel and an extracellular funnel that is filled with a small helical domain, called the plug. During initiation of translocation, a ribosome-nascent chain complex binds to the SecY (or Sec61) complex, resulting in insertion of the nascent chain. However, the mechanism of channel opening during translocation is unclear. Here we have addressed this question by determining structures of inactive and active ribosome-channel complexes with cryo-electron microscopy. Non-translating ribosome-SecY channel complexes derived from Methanocaldococcus jannaschii or Escherichia coli show the channel in its closed state, and indicate that ribosome binding per se causes only minor changes. The structure of an active E. coli ribosome-channel complex demonstrates that the nascent chain opens the channel, causing mostly rigid body movements of the amino- and carboxy-terminal halves of SecY. In this early translocation intermediate, the polypeptide inserts as a loop into the SecY channel with the hydrophobic signal sequence intercalated into the open lateral gate. The nascent chain also forms a loop on the cytoplasmic surface of SecY rather than entering the channel directly. | |||||||||
履歴 |
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構造の表示
ムービー |
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構造ビューア | 分子: ![]() ![]() |
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PDBx/mmCIF形式 | ![]() | 4.2 MB | 表示 | ![]() |
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-関連構造データ
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集合体
登録構造単位 | ![]()
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要素
-Preprotein translocase subunit ... , 2種, 2分子 A7A8
#1: タンパク質 | 分子量: 7701.266 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() プラスミド: pBAD22 / 発現宿主: ![]() ![]() |
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#2: タンパク質 | 分子量: 5853.852 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() プラスミド: pBAD22 / 発現宿主: ![]() ![]() |
+50S ribosomal protein ... , 34種, 35分子 AfAQASATAUAWA5AKAAAaABAbACADAdAEAeAFAgAHAhAIAiAJAjAkALAMANAO...
-タンパク質 , 1種, 1分子 AX
#9: タンパク質 | 分子量: 47482.891 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() プラスミド: pBAD22 / 発現宿主: ![]() ![]() |
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-RNA鎖 , 4種, 4分子 B1B2A1A3
#10: RNA鎖 | 分子量: 24880.865 Da / 分子数: 1 / Fragment: SEE REMARK 999 / 由来タイプ: 天然 由来: (天然) ![]() ![]() |
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#11: RNA鎖 | 分子量: 485455.500 Da / 分子数: 1 / Fragment: SEE REMARK 999 / 由来タイプ: 天然 由来: (天然) ![]() ![]() |
#38: RNA鎖 | 分子量: 990820.375 Da / 分子数: 1 / Fragment: SEE REMARK 999 / 由来タイプ: 天然 由来: (天然) ![]() ![]() |
#39: RNA鎖 | 分子量: 40689.262 Da / 分子数: 1 / Fragment: SEE REMARK 999 / 由来タイプ: 天然 由来: (天然) ![]() ![]() |
+30S ribosomal protein ... , 26種, 27分子 B3AGBABBBCBDBEBFBGBHBIBJBKBLBMBNBOBPBQBRBSBTBUBVBWBXBY
-詳細
配列の詳細 | RIBOSOME COORDINATE |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 / 使用した結晶の数: 1 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
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試料調製
構成要素 |
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分子量 | 値: 2.6 MDa / 実験値: NO | |||||||||||||||||||||||||
緩衝液 | 名称: 100 mM NH4Cl, 30 mM MgCl2, 20 mM HEPES-KOH, 6 mM beta-mercaptoethanol, 0.1% DDM pH: 7.5 詳細: 100 mM NH4Cl, 30 mM MgCl2, 20 mM HEPES-KOH, 6 mM beta-mercaptoethanol, 0.1% DDM | |||||||||||||||||||||||||
試料 | 濃度: 2 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | |||||||||||||||||||||||||
試料支持 | 詳細: Quantifoil 400 mesh 2/1 Cu grids, air glow discharged, and 400 mesh Cu grids with a thin continuous carbon foil | |||||||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK III / 凍結剤: ETHANE / Temp: 77 K / 湿度: 100 % 詳細: Blot 1-2 seconds before plunging into liquid ethane (FEI VITROBOT MARK III). |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Tecnai F20 / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TECNAI F20 / 日付: 2008年4月10日 / 詳細: Low dose imaging, data collected manually |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 50000 X / 倍率(補正後): 51000 X / 最大 デフォーカス(公称値): 2500 nm / 最小 デフォーカス(公称値): 1000 nm / Cs: 2 mm |
試料ホルダ | 試料ホルダーモデル: GATAN LIQUID NITROGEN / 資料ホルダタイプ: Oxford holder / 温度: 93 K |
撮影 | 電子線照射量: 20 e/Å2 / フィルム・検出器のモデル: KODAK SO-163 FILM / 詳細: Kodak SO163 film |
画像スキャン | デジタル画像の数: 217 |
放射波長 | 相対比: 1 |
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解析
EMソフトウェア |
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CTF補正 | 詳細: per micrograph | ||||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C1 (非対称) | ||||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 手法: projection matching / 解像度: 9 Å / 解像度の算出法: FSC 0.5 CUT-OFF / 粒子像の数: 37000 / ピクセルサイズ(公称値): 2.73 Å / ピクセルサイズ(実測値): 2.73 Å 詳細: Final map calculated as an average of 6 different refinements in EMAN2 with different parameters. Resolution method was FSC at 0.5 cut-off using a comparison between experimental map and a ...詳細: Final map calculated as an average of 6 different refinements in EMAN2 with different parameters. Resolution method was FSC at 0.5 cut-off using a comparison between experimental map and a map of the docked ribosomal models calculated at 7 Angstrom resolution with EMAN. クラス平均像の数: 3800 / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||||||||
原子モデル構築 |
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原子モデル構築 | Source name: PDB / タイプ: experimental model
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精密化ステップ | サイクル: LAST
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