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Identifying and characterizing the most significant β-glucosidase of the novel species Aspergillus saccharolyticus

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Abstract:

The newly discovered fungal species Aspergillus saccharolyticus was found to produce a culture broth rich in β-glucosidase activity. In this present work, the main β-glucosidase of A. saccharolyticus responsible for the efficient hydrolytic activity was identified, isolated, and characterized. Ion exchange chromatography was used to fractionate the culture broth, yielding fractions with high β-glucosidase activity and only 1 visible band on an SDS–PAGE gel. Mass spectrometry analysis of this band gave peptide matches to β-glucosidases from aspergilli. Through a polymerase chain reaction approach using degenerate primers and genome walking, a 2919 bp sequence encoding the 860 amino acid BGL1 polypeptide was determined. BGL1 of A. saccharolyticus has 91% and 82% identity with BGL1 from Aspergillus aculeatus and BGL1 from Aspergillus niger, respectively, both belonging to Glycoside Hydrolase family 3. Homology modeling studies suggested β-glucosidase activity with preserved retaining mechanism and a wider catalytic pocket compared with other β-glucosidases. The bgl1 gene was heterologously expressed in Trichoderma reesei QM6a, purified, and characterized by enzyme kinetics studies. The enzyme can hydrolyze cellobiose, p-nitrophenyl-β-d-glucoside, and cellodextrins. The enzyme showed good thermostability, was stable at 50 °C, and at 60 °C it had a half-life of approximately 6 h.

Keywords: Aspergillus saccharolyticus; biomass hydrolysis; caractérisation d’enzyme; enzyme characterization; hydrolyse de la biomasse; thermostability; thermostabilité; β-glucosidase

Document Type: Research Article

DOI: http://dx.doi.org/10.1139/w2012-076

Affiliations: 1: Section for Sustainable Biotechnology, Aalborg University Copenhagen, A.C. Meyers Vænge 15, 2450 København SV, Denmark. 2: MAX-lab, Lund University, Box 118, S-221 00 Lund, Sweden. 3: Chemical and Biological Process Development Group, Energy and Environment Directorate, Pacific Northwest National Laboratory, Richland, WA 99354, USA.

Publication date: September 1, 2012

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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