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

Sorensen, Annette; Ahring, Birgitte K.; Luebeck, Mette; Ubhayasekera, Wimal LU ; Bruno, Kenneth S.; Culley, David E. and Luebeck, Peter S. (2012) In Canadian Journal of Microbiology 58(9). p.1035-1046
Abstract
The newly discovered fungal species Aspergillus saccharolyticus was found to produce a culture broth rich in beta-glucosidase activity. In this present work, the main beta-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 beta-glucosidase activity and only 1 visible band on an SDS-PAGE gel. Mass spectrometry analysis of this band gave peptide matches to beta-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... (More)
The newly discovered fungal species Aspergillus saccharolyticus was found to produce a culture broth rich in beta-glucosidase activity. In this present work, the main beta-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 beta-glucosidase activity and only 1 visible band on an SDS-PAGE gel. Mass spectrometry analysis of this band gave peptide matches to beta-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 beta-glucosidase activity with preserved retaining mechanism and a wider catalytic pocket compared with other beta-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-beta-D-glucoside, and cellodextrins. The enzyme showed good thermostability, was stable at 50 degrees C, and at 60 degrees C it had a half-life of approximately 6 h. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
beta-glucosidase, Aspergillus saccharolyticus, enzyme characterization, thermostability, biomass hydrolysis
in
Canadian Journal of Microbiology
volume
58
issue
9
pages
1035 - 1046
publisher
National Research Council Canada
external identifiers
  • wos:000308368400002
ISSN
0008-4166
DOI
10.1139/W2012-076
language
English
LU publication?
yes
id
91e8e1b5-a2ac-4e60-ac0d-b873888baad0 (old id 3146811)
date added to LUP
2012-11-26 09:47:30
date last changed
2016-04-15 22:00:27
@article{91e8e1b5-a2ac-4e60-ac0d-b873888baad0,
  abstract     = {The newly discovered fungal species Aspergillus saccharolyticus was found to produce a culture broth rich in beta-glucosidase activity. In this present work, the main beta-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 beta-glucosidase activity and only 1 visible band on an SDS-PAGE gel. Mass spectrometry analysis of this band gave peptide matches to beta-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 beta-glucosidase activity with preserved retaining mechanism and a wider catalytic pocket compared with other beta-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-beta-D-glucoside, and cellodextrins. The enzyme showed good thermostability, was stable at 50 degrees C, and at 60 degrees C it had a half-life of approximately 6 h.},
  author       = {Sorensen, Annette and Ahring, Birgitte K. and Luebeck, Mette and Ubhayasekera, Wimal and Bruno, Kenneth S. and Culley, David E. and Luebeck, Peter S.},
  issn         = {0008-4166},
  keyword      = {beta-glucosidase,Aspergillus saccharolyticus,enzyme characterization,thermostability,biomass hydrolysis},
  language     = {eng},
  number       = {9},
  pages        = {1035--1046},
  publisher    = {National Research Council Canada},
  series       = {Canadian Journal of Microbiology},
  title        = {Identifying and characterizing the most significant beta-glucosidase of the novel species Aspergillus saccharolyticus},
  url          = {http://dx.doi.org/10.1139/W2012-076},
  volume       = {58},
  year         = {2012},
}