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GH53 Endo-Beta-1,4-Galactanase from a Newly Isolated Bacillus licheniformis CBMAI 1609 as an Enzymatic Cocktail Supplement for Biomass Saccharification

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Abstract

Galactanases (endo-β-1,4-galactanases—EC 3.2.1.89) catalyze the hydrolysis of β-1,4 galactosidic bonds in arabinogalactan and galactan side chains found in type I rhamnogalacturan. The aim of this work was to understand the catalytic function, biophysical properties, and use of a recombinant GH53 endo-beta-1,4-galactanase for commercial cocktail supplementation. The nucleotide sequence of the endo-β-1,4-galactanase from Bacillus licheniformis CBMAI 1609 (Bl1609Gal) was cloned and expressed in Escherichia coli, and the biochemical and biophysical properties of the enzyme were characterized. The optimum pH range and temperature of Bl1609Gal activity were 6.5–8 and 40 °C, respectively. Furthermore, Bl1609Gal showed remarkable pH stability, retaining more than 75 % activity even after 24 h of incubation at pH 4–10. The enzyme was thermostable, retaining nearly 100 % activity after 1-h incubation at pH 7.0 at 25–45 °C. The enzymatic efficiency (K cat /K m ) against potato galactan under optimum conditions was 241.2 s−1 mg−1 mL. Capillary zone electrophoresis demonstrated that the pattern of galactan hydrolysis by Bl1609Gal was consistent with that of endogalactanases. Supplementation of the commercial cocktail ACCELLERASE®1500 with recombinant Bl1609Gal increased hydrolysis of pretreated sugarcane bagasse by 25 %.

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Acknowledgments

This work was supported by CNPq (481666/2012-5), CAPES, and FAPESP (2013/14454-3) research grants. We also thank the Laboratório de Análises de Macromoléculas (LAM) at Centro Nacional de Pesquisa em Energia e Materiais (CNPEM) for their support on spectroscopic experiments.

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Authors and Affiliations

  1. Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil

    Evandro Antonio de Lima, Carla Botelho Machado, Letícia Maria Zanphorlin, Richard John Ward & Roberto Ruller

  2. Faculdade de Engenharia de Alimentos, Universidade Estadual de Campinas, Campinas, Brazil

    Evandro Antonio de Lima & Hélia Harumi Sato

  3. Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, São Paulo, Brazil

    Richard John Ward

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  1. Evandro Antonio de Lima
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  2. Carla Botelho Machado
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Correspondence to Roberto Ruller.

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de Lima, E.A., Machado, C.B., Zanphorlin, L.M. et al. GH53 Endo-Beta-1,4-Galactanase from a Newly Isolated Bacillus licheniformis CBMAI 1609 as an Enzymatic Cocktail Supplement for Biomass Saccharification. Appl Biochem Biotechnol 179, 415–426 (2016). https://doi.org/10.1007/s12010-016-2003-1

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