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データを開く
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基本情報
登録情報 | データベース: PDB / ID: 2hil | |||||||||
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タイトル | Structure of the Neisseria gonorrhoeae Type IV pilus filament from x-ray crystallography and electron cryomicroscopy | |||||||||
![]() | Fimbrial protein | |||||||||
![]() | CELL ADHESION / TYPE IV PILI / virulence factors / DNA binding protein / natural transformation / antigenic variation | |||||||||
機能・相同性 | ![]() protein secretion by the type II secretion system / type II protein secretion system complex / pilus / cell adhesion / membrane 類似検索 - 分子機能 | |||||||||
生物種 | ![]() | |||||||||
手法 | 電子顕微鏡法 / らせん対称体再構成法 / クライオ電子顕微鏡法 / 解像度: 12.5 Å | |||||||||
![]() | Craig, L. / Volkmann, N. / Egelman, E.H. / Tainer, J.A. | |||||||||
![]() | ![]() タイトル: Type IV pilus structure by cryo-electron microscopy and crystallography: implications for pilus assembly and functions. 著者: Lisa Craig / Niels Volkmann / Andrew S Arvai / Michael E Pique / Mark Yeager / Edward H Egelman / John A Tainer / ![]() 要旨: Type IV pili (T4P) are long, thin, flexible filaments on bacteria that undergo assembly-disassembly from inner membrane pilin subunits and exhibit astonishing multifunctionality. Neisseria ...Type IV pili (T4P) are long, thin, flexible filaments on bacteria that undergo assembly-disassembly from inner membrane pilin subunits and exhibit astonishing multifunctionality. Neisseria gonorrhoeae (gonococcal or GC) T4P are prototypic virulence factors and immune targets for increasingly antibiotic-resistant human pathogens, yet detailed structures are unavailable for any T4P. Here, we determined a detailed experimental GC-T4P structure by quantitative fitting of a 2.3 A full-length pilin crystal structure into a 12.5 A resolution native GC-T4P reconstruction solved by cryo-electron microscopy (cryo-EM) and iterative helical real space reconstruction. Spiraling three-helix bundles form the filament core, anchor the globular heads, and provide strength and flexibility. Protruding hypervariable loops and posttranslational modifications in the globular head shield conserved functional residues in pronounced grooves, creating a surprisingly corrugated pilus surface. These results clarify T4P multifunctionality and assembly-disassembly while suggesting unified assembly mechanisms for T4P, archaeal flagella, and type II secretion system filaments. | |||||||||
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 | ![]() | 536.5 KB | 表示 | ![]() |
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-関連構造データ
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リンク
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集合体
登録構造単位 | ![]()
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要素
#1: タンパク質 | 分子量: 17210.494 Da / 分子数: 18 / 由来タイプ: 天然 / 由来: (天然) ![]() #2: 多糖 | alpha-D-galactopyranose-(1-3)-2,4-bisacetamido-2,4-dideoxy-beta-D-glucopyranose #3: 化合物 | ChemComp-OPE / 配列の詳細 | ACCORDING TO AUTHORS THE RESIDUES HAVE BEEN CONFIRMED BY SEQUENCING THE PILE GENE THAT ENCODES THE ...ACCORDING TO AUTHORS THE RESIDUES HAVE BEEN CONFIRMED BY SEQUENCING | |
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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: FILAMENT / 3次元再構成法: らせん対称体再構成法 |
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試料調製
構成要素 | 名称: N. gonorrhoeae Type IV pili / タイプ: COMPLEX 詳細: native pili were sheared from N. gonorrhoeae cells and purified by successive rounds of precipitation in neutral pH buffer, which causes the filaments to aggregate, followed by resuspension in high pH buffer |
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緩衝液 | 名称: 50 mM CHES / pH: 9.5 / 詳細: 50 mM CHES |
試料 | 濃度: 0.1 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 holey carbon grids, glow discharged with amyl amine |
急速凍結 | 装置: FEI VITROBOT MARK I / 凍結剤: ETHANE 詳細: 5 ul of sample were applied to grids for 1 minute, blotted for 2.5 seconds then plunge-frozen in liquid ethane using an FEI Vitrobot |
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電子顕微鏡撮影
顕微鏡 | モデル: FEI/PHILIPS CM200FEG / 日付: 2004年6月23日 |
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電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 50000 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): 1100 nm |
試料ホルダ | 温度: 95 K / 傾斜角・最大: 0 ° / 傾斜角・最小: 0 ° |
撮影 | 電子線照射量: 10 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: 12 |
放射 | プロトコル: SINGLE WAVELENGTH / 単色(M)・ラウエ(L): M |
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: Wiener filter | ||||||||||||
3次元再構成 | 手法: IHRSR / 解像度: 12.5 Å / 粒子像の数: 25000 / ピクセルサイズ(公称値): 2.23676682 Å / ピクセルサイズ(実測値): 2.23676682 Å / 対称性のタイプ: HELICAL | ||||||||||||
原子モデル構築 | プロトコル: RIGID BODY FIT / 空間: REAL / Target criteria: real-space density correlation coefficient / 詳細: METHOD--CoAn REFINEMENT PROTOCOL--rigid body | ||||||||||||
原子モデル構築 | PDB-ID: 2HI2 | ||||||||||||
精密化 | 最高解像度: 12.5 Å 詳細: The GC pilus filament structure was built by computationally fitting a single GC pilin subunit (2HI2.pdb) into a cryoEM density map of the pilus filament (EMDB code xxx) using the program ...詳細: The GC pilus filament structure was built by computationally fitting a single GC pilin subunit (2HI2.pdb) into a cryoEM density map of the pilus filament (EMDB code xxx) using the program CoAn (Volkmann and Hanein, J. Struct. Biol., 125, 176-184, 1999), and then applying the symmetry operators defined by the cryoEM reconstruction (10.5 Angstrom rise and a 100.8 degree azimuthal rotation, Craig et al., Mol. Cell xxx, yyy, 2006) to the subunit. | ||||||||||||
精密化ステップ | サイクル: LAST / 最高解像度: 12.5 Å
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NMR software | 名称: CoAn / 分類: 精密化 | ||||||||||||
精密化 | ソフトェア番号: 1 詳細: The GC pilus filament structure was built by computationally fitting a single GC pilin subunit (2HI2.pdb) into a cryoEM density map of the pilus filament (EMDB code xxx) using the program ...詳細: The GC pilus filament structure was built by computationally fitting a single GC pilin subunit (2HI2.pdb) into a cryoEM density map of the pilus filament (EMDB code xxx) using the program CoAn (Volkmann and Hanein, J. Struct. Biol., 125, 176-184, 1999), and then applying the symmetry operators defined by the cryoEM reconstruction (10.5 Angstrom rise and a 100.8 degree azimuthal rotation, Craig et al., Mol. Cell xxx, yyy, 2006) to the subunit. |