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EcXyl43 β-xylosidase: molecular modeling, activity on natural and artificial substrates, and synergism with endoxylanases for lignocellulose deconstruction

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Abstract

Biomass hydrolysis constitutes a bottleneck for the biotransformation of lignocellulosic residues into bioethanol and high-value products. The efficient deconstruction of polysaccharides to fermentable sugars requires multiple enzymes acting concertedly. GH43 β-xylosidases are among the most interesting enzymes involved in hemicellulose deconstruction into xylose. In this work, the structural and functional properties of β-xylosidase EcXyl43 from Enterobacter sp. were thoroughly characterized. Molecular modeling suggested a 3D structure formed by a conserved N-terminal catalytic domain linked to an ancillary C-terminal domain. Both domains resulted essential for enzymatic activity, and the role of critical residues, from the catalytic and the ancillary modules, was confirmed by mutagenesis. EcXyl43 presented β-xylosidase activity towards natural and artificial substrates while arabinofuranosidase activity was only detected on nitrophenyl α-L-arabinofuranoside (pNPA). It hydrolyzed xylobiose and purified xylooligosaccharides (XOS), up to degree of polymerization 6, with higher activity towards longer XOS. Low levels of activity on commercial xylan were also observed, mainly on the soluble fraction. The addition of EcXyl43 to GH10 and GH11 endoxylanases increased the release of xylose from xylan and pre-treated wheat straw. Additionally, EcXyl43 exhibited high efficiency and thermal stability under its optimal conditions (40 °C, pH 6.5), with a half-life of 58 h. Therefore, this enzyme could be a suitable additive for hemicellulases in long-term hydrolysis reactions. Because of its moderate inhibition by monomeric sugars but its high inhibition by ethanol, EcXyl43 could be particularly more useful in separate hydrolysis and fermentation (SHF) than in simultaneous saccharification and co-fermentation (SSCF) or consolidated bioprocessing (CBP).

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Acknowledgments

Authors thank MINCYT and CONICET for financial support. OO and FP hold post-doctoral and doctoral fellowships from the Argentinean National Council of Research (CONICET). SG holds an INTA doctoral fellowship. EC, PT, and MC acknowledge CONICET as Research Career members and RM belongs to National Institute of Agriculture Technology (INTA). Pre-treated wheat straw was gently supplied by Dr. Mercedes Ballesteros from CIEMAT, Spain. We thank Igor Polikarpov from USP, Brazil, for critical reading of the manuscript.

Funding

This study was funded by projects MINCYT PICT2014-0791 and CONICET PIP2015-11220150100121CO.

Author information

Authors and Affiliations

  1. Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (INTA), Dr. N. Repetto y Los Reseros s/n, Hurlingham (1686), Buenos Aires, Argentina

    Ornella M. Ontañon, Rubén Marrero Díaz de Villegas, Florencia E. Piccinni, Paola M. Talia & Eleonora Campos

  2. Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Buenos Aires, Argentina

    Ornella M. Ontañon, Florencia E. Piccinni, Paola M. Talia, María L. Cerutti & Eleonora Campos

  3. Instituto de Suelos, CIRN, Instituto Nacional de Tecnología Agropecuaria (INTA), Buenos Aires, Argentina

    Silvina Ghio

  4. Fundación Instituto Leloir, IIBBA-CONICET, Avenida Patricias Argentinas 435, C1405BWE, Buenos Aires, Argentina

    María L. Cerutti

Authors
  1. Ornella M. Ontañon
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  2. Silvina Ghio
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  3. Rubén Marrero Díaz de Villegas
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  4. Florencia E. Piccinni
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  5. Paola M. Talia
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  6. María L. Cerutti
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  7. Eleonora Campos
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Corresponding author

Correspondence to Eleonora Campos.

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Conflict of interest

Ornella Ontañon declares that she has no conflict of interest. Silvina Ghio declares that she has no conflict of interest. Rubén Marrero Díaz de Villegas declares that he has no conflict of interest. Florencia E Piccinni declares that she has no conflict of interest. Paola M Talia declares that she has no conflict of interest. María L Cerutti declares that she has no conflict of interest. Eleonora Campos declares that she has no conflict of interest.

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Ontañon, O.M., Ghio, S., Marrero Díaz de Villegas, R. et al. EcXyl43 β-xylosidase: molecular modeling, activity on natural and artificial substrates, and synergism with endoxylanases for lignocellulose deconstruction. Appl Microbiol Biotechnol 102, 6959–6971 (2018). https://doi.org/10.1007/s00253-018-9138-7

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  • DOI: https://doi.org/10.1007/s00253-018-9138-7

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