A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins
| Autor: | Dritan Siliqi, Alejandra Hernández-Santoyo, Yusvel Sierra-Gómez, Patricia Cano-Sánchez, Annia Rodríguez-Hernández, Homero Gómez-Velasco, Adela Rodríguez-Romero |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Agave tequilana Molecular Dynamics Simulation Biochemistry class I chitinases 03 medical and health sciences 0302 clinical medicine food Agave Coumarins Escherichia coli Glycoside hydrolase Homology modeling Molecular Biology binding energetics Binding Sites Molecular mass biology antifungal proteins Chemistry SAXS models Chitinases Temperature Isothermal titration calorimetry Cell Biology biology.organism_classification food.food 030104 developmental biology Docking (molecular) 030220 oncology & carcinogenesis Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Chitinase biology.protein Thermodynamics Protein Binding |
| Zdroj: | The FEBS journal 286 (2019): 4778–4796. doi:10.1111/febs.14993 info:cnr-pdr/source/autori:Sierra-Gomez, Yusvel; Rodriguez-Hernandez, Annia; Cano-Sanchez, Patricia; Gomez-Velasco, Homero; Hernandez-Santoyo, Alejandra; Siliqi, Dritan; Rodriguez-Romero, Adela/titolo:A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins/doi:10.1111%2Ffebs.14993/rivista:The FEBS journal (Print)/anno:2019/pagina_da:4778/pagina_a:4796/intervallo_pagine:4778–4796/volume:286 |
| DOI: | 10.1111/febs.14993 |
| Popis: | 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)6, 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). |
| Databáze: | OpenAIRE |
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