SASDB75: Escherichia coli TraE protein: A VirB8 homolog from plasmid pKM101

基本情報

• 登録情報: データベース: SASBDB / ID: SASDB75

試料

Escherichia coli TraE protein: A VirB8 homolog from plasmid pKM101

• Escherichia coli TraE protein (VirB8 homolog) (protein), TraE, Escherichia coli

• 機能・相同性: Type IV secretion system protein VirB8/PtlE / Bacterial virulence protein VirB8 / VirB8 protein / protein secretion by the type IV secretion system / NTF2-like domain superfamily / membrane / TraE protein
• 生物種: Escherichia coli (大腸菌)
• 引用: ジャーナル: Proc Natl Acad Sci U S A / 年: 2018; タイトル: VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD.; 著者: Bastien Casu / Charline Mary / Aleksandr Sverzhinsky / Aurélien Fouillen / Antonio Nanci / Christian Baron / ; 要旨: Type IV secretion systems (T4SSs) are multiprotein assemblies that translocate macromolecules across the cell envelope of bacteria. X-ray crystallographic and electron microscopy (EM) analyses have increasingly provided structural information on individual T4SS components and on the entire complex. As of now, relatively little information has been available on the exact localization of the inner membrane-bound T4SS components, notably the mostly periplasmic VirB8 protein and the very hydrophobic VirB6 protein. We show here that the membrane-bound, full-length version of the VirB8 homolog TraE from the plasmid pKM101 secretion system forms a high-molecular-mass complex that is distinct from the previously characterized periplasmic portion of the protein that forms dimers. Full-length TraE was extracted from the membranes with detergents, and analysis by size-exclusion chromatography, cross-linking, and size exclusion chromatography (SEC) multiangle light scattering (MALS) shows that it forms a high-molecular-mass complex. EM and small-angle X-ray scattering (SAXS) analysis demonstrate that full-length TraE forms a hexameric complex with a central pore. We also overproduced and purified the VirB6 homolog TraD and show by cross-linking, SEC, and EM that it binds to TraE. Our results suggest that TraE and TraD interact at the substrate translocation pore of the secretion system.

登録者

• Bastien Casu (Département de biochimie, Université de Montréal, Montréal, Canada)

モデル

• モデル #557: ; タイプ: dummy / ダミー原子の半径: 1.90 A / 対称性: P6 / カイ2乗値: 1.16 / P-value: 0.100000; Omokage検索でこの集合体の類似形状データを探す (詳細)

試料

• 試料: 名称: Escherichia coli TraE protein: A VirB8 homolog from plasmid pKM101
• バッファ: 名称: 50 mM sodium phosphate 300 mM NaCl 40 mM imidazole 0.15 % octyl glucose neopentyl glycol (OGNG); 濃度: 50.00 mM / pH: 7.4 ; 組成: 300 mM NaCl, 40 mM imidazole, 0.15 % octyl glucose neopentyl glycol (OGNG)

要素 #358

名称: TraE / タイプ: protein / 記述: Escherichia coli TraE protein (VirB8 homolog) / 分子量: 28.537 / 分子数: 6 / 由来: Escherichia coli / 参照: UniProt: Q46703
配列:

MGSSHHHHHH ENLYFQGGTK ANKKTGLTRE AIKEFNESRK GLEVDLMDEV LKSRRTAWMV ATGSAVVTVF ALSLVGYVVH KYSQPIPAHL LTLNEATHEV QQVKLTRDQT SYGDEIDKFW LTQYVIHRES YDFYSVQVDY TAVGLMSTPN VAESYQSKFK GRNGLDKVLG DSETTRVKIN SVILDKPHGV ATIRFTTVRR VRSNPVDDQP QRWIAIMGYE YKSLAMNAEQ RYVNPLGFRV TSYRVNPEVN

実験情報

• ビーム: 設備名称: Cornell High Energy Synchrotron Source (CHESS) G1; 地域: Ithaca, NY / 国: USA / 線源: X-ray synchrotron
• 検出器: 名称: Finger Lakes CCD / タイプ: CCD

スキャン

タイトル: TraE protein / 測定日: 2016年6月2日 / セル温度: 20 °C / 照射時間: 2 sec. / フレーム数: 70 / 単位: 1/nm /

	Min	Max
Q	0.0915	2.6113

距離分布関数 P(R)

ソフトウェア P(R): GNOM 4.6 / ポイント数: 441 /

	Min	Max
Q	0.009152	0.2606
P(R) point	1	441
R	0	137

結果

カーブのタイプ: single_conc
コメント: The MW of the TraE/OGNG complex was additionally confirmed using SEC-MALLS (248kDa).

	実験値	Porod
分子量	264.2 kDa	223 kDa
体積	-	360 nm3

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	0.0094	3.0E-5	0.0094	6.0E-5
慣性半径, Rg	4.44 nm	0.013	4.44 nm	0.13

	Min	Max
D	-	13.7
Guinier point	10	35