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One-Step Immobilization and Stabilization of a Recombinant Enterococcus faecium DBFIQ E36 l-Arabinose Isomerase for d-Tagatose Synthesis

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Abstract

A recombinant l-arabinose isomerase from Enterococcus faecium DBFIQ E36 was immobilized onto multifunctional epoxide supports by chemical adsorption and onto a chelate-activated support via polyhistidine-tag, located on the N-terminal (N-His-L-AI) or on the C-terminal (C-His-L-AI) sequence, followed by covalent bonding between the enzyme and the support. The results were compared to reversible L-AI immobilization by adsorption onto charged agarose supports with improved stability. All the derivatives presented immobilization yields of above 75%. The ionic interaction established between agarose gels containing monoaminoethyl-N-aminoethyl structures (MANAE) and the enzyme was the most suitable strategy for L-AI immobilization in comparison to the chelate-activated agarose. In addition, the immobilized biocatalysts by ionic interaction in MANAE showed to be the most stable, retaining up to 100% of enzyme activity for 60 min at 60 °C and with Km values of 28 and 218 mM for MANAE-N-His-L-AI and MANAE-C-His-L-AI, respectively.

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Funding

This study received financial support from the Brazilian Research Agencies CAPES, CNPq, and FUNCAP. This work was partially sponsored by funds from the project Argentina–Brazil Bilateral Cooperation Program BR/12/06 MINCyT-CAPES 2012 (Buenos Aires, Argentina).

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Authors and Affiliations

  1. Department of Chemical Engineering, Federal University of Ceará, Campus do Pici, BL 709, Fortaleza, CE, Brazil

    Marylane de Sousa & Luciana R. B. Gonçalves

  2. Department of Biology, Federal University of Ceará, Campus do Pici, BL 909, Fortaleza, CE, Brazil

    Vânia M. M. Melo & Denise C. Hissa

  3. Food and Biotechnology Engineering Group, Institute of Technological Development for the Chemical Industry, National University of the Litoral (UNL), National Council of Scientific and Technical Research (CONICET), RN 168 Km 472 “Paraje El Pozo” S / N, Santa Fe, Argentina

    Ricardo M. Manzo & Enrique J. Mammarella

  4. Department of Infectious Diseases, King’s College London, London, WC2R 2LS, UK

    André Saraiva Leão Marcelo Antunes

  5. Center for Biological Research, CIB, Higher Council for Scientific Research, CSIC, C / Ramiro de Maeztu, 9, Madrid, Spain

    José L. García

  6. Department of Food Biotechnology and Microbiology, Institute of Research in Food Sciences, CIAL, Higher Council for Scientific Research, CSIC, C / Nicolás Cabrera 9, UAM Campus, Madrid, Spain

    Benevides C. Pessela

  7. Department of Engineering and Technology, Polytechnic Institute of Sciences and Technology, Av. Luanda Sul, Rua Lateral Via S10, Talatona, Luanda, Angola

    Benevides C. Pessela

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  1. Marylane de Sousa
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  2. Vânia M. M. Melo
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  3. Denise C. Hissa
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  5. Enrique J. Mammarella
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  6. André Saraiva Leão Marcelo Antunes
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  8. Benevides C. Pessela
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  9. Luciana R. B. Gonçalves
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Correspondence to Benevides C. Pessela or Luciana R. B. Gonçalves.

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de Sousa, M., Melo, V.M.M., Hissa, D.C. et al. One-Step Immobilization and Stabilization of a Recombinant Enterococcus faecium DBFIQ E36 l-Arabinose Isomerase for d-Tagatose Synthesis. Appl Biochem Biotechnol 188, 310–325 (2019). https://doi.org/10.1007/s12010-018-2905-1

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