In silico screening and experimental analysis of family GH11 xylanases for applications under conditions of alkaline pH
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otechnology for Biofuels Open Access
RESEARCH
In silico screening and experimental analysis of family GH11 xylanases for applications under conditions of alkaline pH and high temperature David Talens‑Perales1, Paloma Sánchez‑Torres1, Julia Marín‑Navarro1,2 and Julio Polaina1*
Abstract Background: Xylanases are one of the most extensively used enzymes for biomass digestion. However, in many instances, their use is limited by poor performance under the conditions of pH and temperature required by the industry. Therefore, the search for xylanases able to function efficiently at alkaline pH and high temperature is an important objective for different processes that use lignocellulosic substrates, such as the production of paper pulp and biofuels. Results: A comprehensive in silico analysis of family GH11 sequences from the CAZY database allowed their phy‑ logenetic classification in a radial cladogram in which sequences of known or presumptive thermophilic and alka‑ lophilic xylanases appeared in three clusters. Eight sequences from these clusters were selected for experimental analysis. The coding DNA was synthesized, cloned and the enzymes were produced in E. coli. Some of these showed high xylanolytic activity at pH values > 8.0 and temperature > 80 °C. The best enzymes corresponding to sequences from Dictyoglomus thermophilum (Xyn5) and Thermobifida fusca (Xyn8). The addition of a carbohydrate-binding mod‑ ule (CBM9) to Xyn5 increased 4 times its activity at 90 °C and pH > 9.0. The combination of Xyn5 and Xyn8 was proved to be efficient for the saccharification of alkali pretreated rice straw, yielding xylose and xylooligosaccharides. Conclusions: This study provides a fruitful approach for the selection of enzymes with suitable properties from the information contained in extensive databases. We have characterized two xylanases able to hydrolyze xylan with high efficiency at pH > 8.0 and temperature > 80 °C. Keywords: Carbohydrate-binding domain, Glycoside hydrolase, Rice straw, Xylose, Xylooligosaccharides Background Xylan, the most abundant type of hemicellulose, is a polysaccharide composed by a linear backbone of β-1, 4-linked xylose units. Together with cellulose and lignin, xylan is one of the main constituents of plant cell walls. *Correspondence: [email protected] 1 Department of Food Biotechnology. Institute of Agrochemistry and Food Technology, Spanish National Research Council (IATA-CSIC), Paterna, Valencia, Spain Full list of author information is available at the end of the article
Xylan degradation into simple sugars, a preliminary step for its conversion into different bioproducts, is carried out by the concerted action of different xylanolytic enzymes [1, 2]. Xylanases produced by many microbial species are an important type of industrial enzymes with multiple applications. They are used as additives to enhance the quality of baked goods [3] and animal feeds [4], as well as to bleach kraft pulp [5, 6]. Enzymatic biotransformation of xylan is limited by different factors such as the natu
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