SASDCK9: Catalytic domain (AC) of B. Pertussis Adenylate Cyclase Toxin (CyaA)

基本情報

• 登録情報: データベース: SASBDB / ID: SASDCK9

試料

Catalytic domain (AC) of B. Pertussis Adenylate Cyclase Toxin (CyaA)

• Bifunctional hemolysin/adenylate cyclase (protein), AC domain from CyaA, Bordetella pertussis (strain Tohama I / ATCC BAA-589 / NCTC 13251)

機能・相同性

分子機能:

calcium- and calmodulin-responsive adenylate cyclase activity / hemolysis in another organism / adenylate cyclase / cAMP biosynthetic process / adenylate cyclase activity / channel activity / toxin activity / positive regulation of cytosolic calcium ion concentration / calmodulin binding / calcium ion binding / host cell plasma membrane / extracellular region / ATP binding / membrane

ドメイン・相同性:

RTX, pore-forming domain / N-terminal domain in RTX protein / Haemolysin-type calcium binding-related / Haemolysin-type calcium binding protein related domain / RTX toxin determinant A / Anthrax toxin, edema factor, central / Anthrax toxin, edema factor, C-terminal / Anthrax toxin, edema factor, central domain superfamily / Anthrax toxin LF subunit / Hemolysin-type calcium-binding conserved site / Hemolysin-type calcium-binding region signature. / RTX calcium-binding nonapeptide repeat / RTX calcium-binding nonapeptide repeat (4 copies) / Serralysin-like metalloprotease, C-terminal

構成要素:

Bifunctional hemolysin/adenylate cyclase

• 生物種: Bordetella pertussis (strain Tohama I / ATCC BAA-589 / NCTC 13251) (百日咳菌)
• 引用: ジャーナル: PLoS Biol / 年: 2017; タイトル: Calmodulin fishing with a structurally disordered bait triggers CyaA catalysis.; 著者: Darragh P O'Brien / Dominique Durand / Alexis Voegele / Véronique Hourdel / Marilyne Davi / Julia Chamot-Rooke / Patrice Vachette / Sébastien Brier / Daniel Ladant / Alexandre Chenal / ; 要旨: Once translocated into the cytosol of target cells, the catalytic domain (AC) of the adenylate cyclase toxin (CyaA), a major virulence factor of Bordetella pertussis, is potently activated by binding calmodulin (CaM) to produce supraphysiological levels of cAMP, inducing cell death. Using a combination of small-angle X-ray scattering (SAXS), hydrogen/deuterium exchange mass spectrometry (HDX-MS), and synchrotron radiation circular dichroism (SR-CD), we show that, in the absence of CaM, AC exhibits significant structural disorder, and a 75-residue-long stretch within AC undergoes a disorder-to-order transition upon CaM binding. Beyond this local folding, CaM binding induces long-range allosteric effects that stabilize the distant catalytic site, whilst preserving catalytic loop flexibility. We propose that the high enzymatic activity of AC is due to a tight balance between the CaM-induced decrease of structural flexibility around the catalytic site and the preservation of catalytic loop flexibility, allowing for fast substrate binding and product release. The CaM-induced dampening of AC conformational disorder is likely relevant to other CaM-activated enzymes.

登録者

• Dominique DURAND (I2BC-CNRS)

モデル

• モデル #1690: ; タイプ: mix / ダミー原子の半径: 1.90 A / カイ2乗値: 2.565; Omokage検索でこの集合体の類似形状データを探す (詳細)
• モデル #1691: ; タイプ: atomic / ダミー原子の半径: 1.90 A / カイ2乗値: 1.358 / P-value: 0.000720; 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: Catalytic domain (AC) of B. Pertussis Adenylate Cyclase Toxin (CyaA)
• バッファ: 名称: 20 mM Hepes, 150 mM NaCl, 4 mM CaCl2 / pH: 7.4

要素 #904

名称: AC domain from CyaA / タイプ: protein / 記述: Bifunctional hemolysin/adenylate cyclase / 分子量: 39.38 / 分子数: 1
由来: Bordetella pertussis (strain Tohama I / ATCC BAA-589 / NCTC 13251)
参照: UniProt: P0DKX7
配列:

MQQSHQAGYA NAADRESGIP AAVLDGIKAV AKEKNATLMF RLVNPHSTSL IAEGVATKGL GVHAKSSDWG LQAGYIPVNP NLSKLFGRAP EVIARADNDV NSSLAHGHTA VDLTLSKERL DYLRQAGLVT GMADGVVASN HAGYEQFEFR VKETSDGRYA VQYRRKGGDD FEAVKVIGNA AGIPLTADID MFAIMPHLSN FRDSARSSVT SGDSVTDYLA RTRRAASEAT GGLDRERIDL LWKIARAGAR SAVGTEARRQ FRYDGDMNIG VITDFELEVR NALNRRAHAV GAQDVVQHGT EQNNPFPEAD EKIFVVSATG ESQMLTRGQL KEYIGQQRGE GYVFYENRAY GVAGKSLFDD GLGA

実験情報

• ビーム: 設備名称: SOLEIL SWING / 地域: Saint-Aubin / 国: France / 線源: X-ray synchrotron / 波長: 0.1 Å / スペクトロメータ・検出器間距離: 1.988 mm
• 検出器: 名称: AVIEX / タイプ: CCD

スキャン

タイトル: Catalytic domain (AC) of B. Pertussis Adenylate Cyclase Toxin (CyaA)
測定日: 2015年6月19日 / 保管温度: 10 °C / セル温度: 15 °C / 照射時間: 1.5 sec. / フレーム数: 250 / 単位: 1/A /

	Min	Max
Q	0.0066	0.3999

距離分布関数 P(R)

ソフトウェア P(R): GNOM 5.0 / ポイント数: 773 /

	Min	Max
Q	0.00663373	0.399925
P(R) point	1	773
R	0	91

結果

カーブのタイプ: sec
コメント: The scattered intensities were displayed on an absolute scale (cm-1) using the scattering of water. Frames were examined individually and 20 identical frames were averaged and further processed. The corresponding concentration was 0.80 g/L. Three independent determinations of the molecular mass were obtained from the value of I(0)/c, where c is the protein concentration, and using the programs SAXSMow2 and ScÅtter3 available at the URLs http://saxs.ifsc.usp.br/ and https://bl1231.als.lbl.gov/scatter/, respectively. The average value is MWexperimental=40.2 ± 1.0 kDa. AC in solution: Top panel: Adjustment of the curve calculated from the crystal structure of AC extracted from the pdb dataset 1YRU (red curve) after missing loop addition using AllosModFoxs against experimental data (blue dots). chi2=2.57 Bottom panel: Adjustment obtained using AllosModFoxs by releasing residues 200-270 comprising helices F through H’ (red curve). Residues 200-270 correspond to the large yellow-light green loop seen in the model displayed on the right. chi2=1.36

	実験値	Standard	Standard error	Porod
分子量	40.2 kDa	38.6 kDa	2	40.1 kDa
体積	-	-	-	60.2 nm3

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	0.0252	0.0002	0.0252	0.0002
慣性半径, Rg	2.62 nm	0.03	2.59 nm	0.03

	Min	Max	Error
D	-	9.1	0.5
Guinier point	1	66	-