SASDDE7: Class I chitinase 1 from Agave tequilana (Chitinase 1, ChiAt1)

基本情報

• 登録情報: データベース: SASBDB / ID: SASDDE7

試料

Class I chitinase 1 from Agave tequilana

• Chitinase 1 (protein), ChiAt1, Agave tequilana

• 生物種: Agave tequilana (植物)
• 引用: ジャーナル: FEBS J / 年: 2019; タイトル: A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins.; 著者: Yusvel Sierra-Gómez / Annia Rodríguez-Hernández / Patricia Cano-Sánchez / Homero Gómez-Velasco / Alejandra Hernández-Santoyo / Dritan Siliqi / Adela Rodríguez-Romero / ; 要旨: Plant chitinases are enzymes that have several functions, including providing protection against pathogens. Agave tequilana is an economically important plant that is poorly studied. Here, we identified a chitinase from short reads of the A. tequilana transcriptome (AtChi1). A second chitinase, differing by only six residues from the first, was isolated from total RNA of plants infected with Fusarium oxysporum (AtChi2). Both enzymes were overexpressed in Escherichia coli and analysis of their sequences indicated that they belong to the class I glycoside hydrolase family19, whose members exhibit two domains: a carbohydrate-binding module and a catalytic domain, connected by a flexible linker. Activity assays and thermal shift experiments demonstrated that the recombinant Agave enzymes are highly thermostable acidic endochitinases with Tm values of 75 °C and 71 °C. Both exhibit a molecular mass close to 32 kDa, as determined by MALDI-TOF, and experimental pIs of 3.7 and 3.9. Coupling small-angle x-ray scattering information with homology modeling and docking simulations allowed us to structurally characterize both chitinases, which notably show different interactions in the binding groove. Even when the six different amino acids are all exposed to solvent in the loops located near the linker and opposite to the binding site, they confer distinct kinetic parameters against colloidal chitin and similar affinity for (GlnNAc) as shown by isothermal titration calorimetry. Interestingly, binding is more enthalpy-driven for AtChi2. Whereas the physiological role of these chitinases remains unknown, we demonstrate that they exhibit important antifungal activity against chitin-rich fungi such as Aspergillus sp. DATABASE: SAXS structural data are available in the SASBDB database with accession numbers SASDDE7 and SASDDA6. ENZYMES: Chitinases (EC3.2.1.14).

登録者

• Yusvel Sierra-Gómez (IQ, UNAM, Mexico)

モデル

• モデル #2037: ; タイプ: dummy / ダミー原子の半径: 1.40 A / カイ2乗値: 1.624 / P-value: 0.216071; 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: Class I chitinase 1 from Agave tequilana / 試料濃度: 0.23 mg/ml
• バッファ: 名称: MES 50 mM / pH: 6

要素 #1052

名称: ChiAt1 / タイプ: protein / 記述: Chitinase 1 / 分子量: 31.753 / 分子数: 1 / 由来: Agave tequilana
配列:

GGAQQCGSQA GGAVCPNGLC CSQFGYCGST SPYCGNGCQS QCGGGSSPTP NPPSGGGGGG GGGGSGVGSI ISSSLFDQIL LHRNDAACPA HGFYTYDAFV AAANAFSGFA TTGDADTQKR EIAAFLAQTS HETTGGWPTA PDGPYSWGYC FLQEQGNPGD YCVPNDQWPC APGKKYYGRG PIQISYNYNY GPCGNAIGSD LLNNPDLVAS DPTVSFKTAL WFWMTPQSPK PSCHDVITGQ WTPSAADQAA GRVPGFGVIT NIINGGVECG HGSDSRDEDR VGFYKRYCDI LGVSYGDNLD CGNQSHF

実験情報

• ビーム: 設備名称: Stanford Synchrotron Radiation Lightsource (SSRL) BL4-2; 地域: Menlo Park, CA / 国: USA / 線源: X-ray synchrotron / 波長: 1.127 Å / スペクトロメータ・検出器間距離: 1.78 mm
• 検出器: 名称: Rayonix MX225-HE

スキャン

タイトル: ChiAt1 / 測定日: 2017年4月26日 / 保管温度: 4 °C / セル温度: 15 °C / 照射時間: 1 sec. / 単位: 1/A /

	Min	Max
Q	0.0132	0.4188

距離分布関数 P(R)

ソフトウェア P(R): GNOM 5.0 / ポイント数: 330 /

	Min	Max
Q	0.0168229	0.318071
P(R) point	1	330
R	0	96.6

結果

カーブのタイプ: single_conc

	実験値	Standard	Standard error	Porod
分子量	31.7 kDa	30.5 kDa	2	32.5 kDa
体積	-	-	-	52 nm3

	P(R)	Guinier	Guinier error
前方散乱 I0	257.2	257.54	1.72
慣性半径, Rg	2.61 nm	2.551 nm	0.029

	Min	Max
D	-	9.66
Guinier point	5	38