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基本情報
登録情報 | データベース: PDB / ID: 2bk2 | ||||||
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タイトル | The prepore structure of pneumolysin, obtained by fitting the alpha carbon trace of perfringolysin O into a cryo-EM map | ||||||
![]() | PERFRINGOLYSIN O | ||||||
![]() | TOXIN / CYTOLYSIS / HEMOLYSIS / THIOL-ACTIVATED CYTOLYSIN / CRYOEM / CYTOLYTIC PROTEIN | ||||||
機能・相同性 | ![]() hemolysis in another organism / cholesterol binding / toxin activity / membrane => GO:0016020 / host cell plasma membrane / extracellular region / 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: 28 Å | ||||||
Model type details | CA ATOMS ONLY, CHAIN A | ||||||
![]() | Tilley, S.J. / Orlova, E.V. / Gilbert, R.J.C. / Andrew, P.W. / Saibil, H.R. | ||||||
![]() | ![]() タイトル: Structural basis of pore formation by the bacterial toxin pneumolysin. 著者: Sarah J Tilley / Elena V Orlova / Robert J C Gilbert / Peter W Andrew / Helen R Saibil / ![]() 要旨: The bacterial toxin pneumolysin is released as a soluble monomer that kills target cells by assembling into large oligomeric rings and forming pores in cholesterol-containing membranes. Using cryo-EM ...The bacterial toxin pneumolysin is released as a soluble monomer that kills target cells by assembling into large oligomeric rings and forming pores in cholesterol-containing membranes. Using cryo-EM and image processing, we have determined the structures of membrane-surface bound (prepore) and inserted-pore oligomer forms, providing a direct observation of the conformational transition into the pore form of a cholesterol-dependent cytolysin. In the pore structure, the domains of the monomer separate and double over into an arch, forming a wall sealing the bilayer around the pore. This transformation is accomplished by substantial refolding of two of the four protein domains along with deformation of the membrane. Extension of protein density into the bilayer supports earlier predictions that the protein inserts beta hairpins into the membrane. With an oligomer size of up to 44 subunits in the pore, this assembly creates a transmembrane channel 260 A in diameter lined by 176 beta strands. #1: ![]() タイトル: Structure of a cholesterol-binding, thiol-activated cytolysin and a model of its membrane form. 著者: J Rossjohn / S C Feil / W J McKinstry / R K Tweten / M W Parker / ![]() 要旨: The mechanisms by which proteins gain entry into membranes is a fundamental problem in biology. Here, we present the first crystal structure of a thiol-activated cytolysin, perfringolysin O, a member ...The mechanisms by which proteins gain entry into membranes is a fundamental problem in biology. Here, we present the first crystal structure of a thiol-activated cytolysin, perfringolysin O, a member of a large family of toxins that kill eukaryotic cells by punching holes in their membranes. The molecule adopts an unusually elongated shape rich in beta sheet. We have used electron microscopy data to construct a detailed model of the membrane channel form of the toxin. The structures reveal a novel mechanism for membrane insertion. Surprisingly, the toxin receptor, cholesterol, appears to play multiple roles: targeting, promotion of oligomerization, triggering a membrane insertion competent form, and stabilizing the membrane pore. | ||||||
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構造の表示
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構造ビューア | 分子: ![]() ![]() |
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-検証レポート
文書・要旨 | ![]() | 646 KB | 表示 | ![]() |
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文書・詳細版 | ![]() | 645.6 KB | 表示 | |
XML形式データ | ![]() | 11.4 KB | 表示 | |
CIF形式データ | ![]() | 15.7 KB | 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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対称性 | 点対称性: (シェーンフリース記号: C31 (31回回転対称)) |
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要素
#1: タンパク質 | 分子量: 50992.805 Da / 分子数: 1 / 断片: RESIDUES 36-500 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 発現宿主: ![]() ![]() |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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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 | 名称: PNEUMOLYSIN / タイプ: COMPLEX 詳細: THE SAMPLE CONSISTS OF PNEUMOLYSIN IN A MEMBRANE-INSERTED PORE STATE |
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緩衝液 | 名称: 8 MM NA2HP04, 1.5MM KH2PO4, 2.5 MM KCL, 0.25 MM NACL pH: 6.95 詳細: 8 MM NA2HP04, 1.5MM KH2PO4, 2.5 MM KCL, 0.25 MM NACL |
試料 | 濃度: 0.05 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: HOLEY CARBON |
急速凍結 | 装置: HOMEMADE PLUNGER / 凍結剤: ETHANE / 詳細: PLUNGED INTO ETHANE |
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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 詳細: SAMPLES WERE MAINTAINED AT LIQUID NITROGEN TEMPERATURES IN THE ELECTRON MICROSCOPE. |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 42000 X / 最大 デフォーカス(公称値): 3200 nm / 最小 デフォーカス(公称値): 1100 nm / Cs: 2 mm |
試料ホルダ | 温度: 100 K / 傾斜角・最大: 0 ° / 傾斜角・最小: 0 ° |
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 |
画像スキャン | デジタル画像の数: 135 |
放射波長 | 相対比: 1 |
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解析
CTF補正 | 詳細: PHASE FLIPPING | ||||||||||||
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対称性 | 点対称性: C31 (31回回転対称) | ||||||||||||
3次元再構成 | 解像度: 28 Å / 粒子像の数: 223 / ピクセルサイズ(公称値): 3.5 Å 詳細: RESIDUES 134-142 ARE MISSING FROM THE SEQUENCE. THE 31-MER CAN BE GENERATED BY APPLYING 31-FOLD ROTATIONAL SYMMETRY 対称性のタイプ: POINT | ||||||||||||
原子モデル構築 | プロトコル: RIGID BODY FIT 詳細: METHOD--THE CRYSTAL STRUCTURE OF PERFRINOGLYSIN O (1PFO, ROSSJOHN ET AL., 1998, CELL 89, 685)WAS PLACED INTO THE CRYO-EM DENSITY MAP (EMD-1106). THE ALPHA CARBON TRACE OF PERFRINGOLYSIN 0 WAS ...詳細: METHOD--THE CRYSTAL STRUCTURE OF PERFRINOGLYSIN O (1PFO, ROSSJOHN ET AL., 1998, CELL 89, 685)WAS PLACED INTO THE CRYO-EM DENSITY MAP (EMD-1106). THE ALPHA CARBON TRACE OF PERFRINGOLYSIN 0 WAS MANUALLY POSITIONED INTO THE CRYO-EM DENSITY CORRESPONDING TO THE POSITION OF ONE SUBUNIT. THE BEST FIT WAS OBTAINED BY SEPARATING THE MONOMER INTO TWO RIGID BODIES- DOMAINS 1-3 (36-390) AND DOMAIN 4 (391-500). THE COMPLETE OLIGOMER (31-MER) WAS GENERATED AND CHECKED FOR CLOSE CONTACTS BOTH BY EYE AND USING THE CCP4 PROGRAM CONTACT. | ||||||||||||
原子モデル構築 | PDB-ID: 1PFO Accession code: 1PFO / Source name: PDB / タイプ: experimental model | ||||||||||||
精密化 | 最高解像度: 28 Å | ||||||||||||
精密化ステップ | サイクル: LAST / 最高解像度: 28 Å
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