+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 8ftf | ||||||
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タイトル | CryoEM strucutre of 33-mer RBM3 of the Salmonella MS-ring | ||||||
要素 | Flagellar M-ring protein | ||||||
キーワード | MOTOR PROTEIN / flagellar / 33-fold / ring-building motif / MS-ring / FliF | ||||||
機能・相同性 | 機能・相同性情報 bacterial-type flagellum basal body, MS ring / cytoskeletal motor activity / bacterial-type flagellum-dependent cell motility / plasma membrane 類似検索 - 分子機能 | ||||||
生物種 | Salmonella enterica subsp. enterica serovar Typhimurium (サルモネラ菌) | ||||||
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: 2.9 Å | ||||||
データ登録者 | Singh, P.S. / Nakagawa, T. / Cecchini, G. / Iverson, T.M. | ||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: PLoS One / 年: 2023 タイトル: CryoEM structure of a post-assembly MS-ring reveals plasticity in stoichiometry and conformation. 著者: Prashant K Singh / Gary Cecchini / Terunaga Nakagawa / T M Iverson / 要旨: The flagellar motor supports bacterial chemotaxis, a process that allows bacteria to move in response to their environment. A central feature of this motor is the MS-ring, which is composed entirely ...The flagellar motor supports bacterial chemotaxis, a process that allows bacteria to move in response to their environment. A central feature of this motor is the MS-ring, which is composed entirely of repeats of the FliF subunit. This MS-ring is critical for the assembly and stability of the flagellar switch and the entire flagellum. Despite multiple independent cryoEM structures of the MS-ring, there remains a debate about the stoichiometry and organization of the ring-building motifs (RBMs). Here, we report the cryoEM structure of a Salmonella MS-ring that was purified from the assembled flagellar switch complex (MSC-ring). We term this the 'post-assembly' state. Using 2D class averages, we show that under these conditions, the post-assembly MS-ring can contain 32, 33, or 34 FliF subunits, with 33 being the most common. RBM3 has a single location with C32, C33, or C34 symmetry. RBM2 is found in two locations with RBM2inner having C21 or C22 symmetry and an RBM2outer-RBM1 having C11 symmetry. Comparison to previously reported structures identifies several differences. Most strikingly, we find that the membrane domain forms 11 regions of discrete density at the base of the structure rather than a contiguous ring, although density could not be unambiguously interpreted. We further find density in some previously unresolved areas, and we assigned amino acids to those regions. Finally, we find differences in interdomain angles in RBM3 that affect the diameter of the ring. Together, these investigations support a model of the flagellum with structural plasticity, which may be important for flagellar assembly and function. | ||||||
履歴 |
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-構造の表示
構造ビューア | 分子: MolmilJmol/JSmol |
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-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 8ftf.cif.gz | 1 MB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb8ftf.ent.gz | 表示 | PDB形式 | |
PDBx/mmJSON形式 | 8ftf.json.gz | ツリー表示 | PDBx/mmJSON形式 | |
その他 | その他のダウンロード |
-検証レポート
文書・要旨 | 8ftf_validation.pdf.gz | 1.5 MB | 表示 | wwPDB検証レポート |
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文書・詳細版 | 8ftf_full_validation.pdf.gz | 1.5 MB | 表示 | |
XML形式データ | 8ftf_validation.xml.gz | 157.7 KB | 表示 | |
CIF形式データ | 8ftf_validation.cif.gz | 194.9 KB | 表示 | |
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/ft/8ftf ftp://data.pdbj.org/pub/pdb/validation_reports/ft/8ftf | HTTPS FTP |
-関連構造データ
関連構造データ | 29425MC 8fteC M: このデータのモデリングに利用したマップデータ C: 同じ文献を引用 (文献) |
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類似構造データ | 類似検索 - 機能・相同性F&H 検索 |
-リンク
-集合体
登録構造単位 |
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1 |
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-要素
#1: タンパク質 | 分子量: 61295.645 Da / 分子数: 33 / 由来タイプ: 組換発現 由来: (組換発現) Salmonella enterica subsp. enterica serovar Typhimurium (サルモネラ菌) 遺伝子: fliF, fla AII.1, fla BI, STM1969 / 発現宿主: Escherichia coli BL21(DE3) (大腸菌) / 参照: UniProt: P15928 |
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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 |
-試料調製
構成要素 | 名称: MS-ring of the flagellar complex / タイプ: COMPLEX 詳細: CryoEM strucutre of the 33-mer RBM3 region of the Salmonella MS-ring Entity ID: all / 由来: RECOMBINANT | ||||||||||||||||||||
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分子量 | 値: 2.5 MDa / 実験値: NO | ||||||||||||||||||||
由来(天然) | 生物種: Salmonella enterica subsp. enterica serovar Typhimurium (サルモネラ菌) | ||||||||||||||||||||
由来(組換発現) | 生物種: Escherichia coli BL21(DE3) (大腸菌) | ||||||||||||||||||||
緩衝液 | pH: 7.5 詳細: 50 mM Tris-HCl pH 7.5, 100 mM NaCl, 0.1 % Ana-grade LDAO | ||||||||||||||||||||
緩衝液成分 |
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試料 | 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | ||||||||||||||||||||
試料支持 | 詳細: Quantifoil grid coated with graphene oxide substrate (Electron Microscopy Sciences) was glow discharged for 5 sec. Purified MS-rings were added to each grid at 4C at 100% humidity. After 30 ...詳細: Quantifoil grid coated with graphene oxide substrate (Electron Microscopy Sciences) was glow discharged for 5 sec. Purified MS-rings were added to each grid at 4C at 100% humidity. After 30 sec of incubation, blotting was performed for 12 sec. The grid was then plunged into liquid ethane using a Vitrobot Mark IV system (Thermo Fisher). グリッドの材料: GRAPHENE OXIDE / グリッドのサイズ: 400 divisions/in. / グリッドのタイプ: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK IV / 凍結剤: ETHANE / 湿度: 100 % / 凍結前の試料温度: 277.15 K 詳細: Purified MS-rings were added to each grid at 277.15K at 100% humidity. After 30 sec of incubation, blotting was performed for 12 sec. The grid was then plunged into liquid ethane using a ...詳細: Purified MS-rings were added to each grid at 277.15K at 100% humidity. After 30 sec of incubation, blotting was performed for 12 sec. The grid was then plunged into liquid ethane using a Vitrobot Mark IV system (Thermo Fisher) |
-電子顕微鏡撮影
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: FIELD EMISSION GUN / 加速電圧: 300 kV / 照射モード: FLOOD BEAM |
電子レンズ | モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 2200 nm / 最小 デフォーカス(公称値): 800 nm |
撮影 | 電子線照射量: 50 e/Å2 フィルム・検出器のモデル: FEI FALCON III (4k x 4k) |
-解析
EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C33 (33回回転対称) | ||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 2.9 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 32042 / 対称性のタイプ: POINT |