title: Identification and characterization of core cellulolytic enzymes from Talaromyces cellulolyticus (formerly Acremonium cellulolyticus) critical for hydrolysis of lignocellulosic biomass.
abstract: BACKGROUND
  Enzymatic hydrolysis of pretreated lignocellulosic biomass is an essential process for the production of fermentable sugars for industrial use. A better understanding of fungal cellulase systems will provide clues for maximizing the hydrolysis of target biomass. Talaromyces cellulolyticus is a promising fungus for cellulase production and efficient biomass hydrolysis. Several cellulolytic enzymes purified from T. cellulolyticus were characterized in earlier studies, but the core enzymes critical for hydrolysis of lignocellulosic biomass remain unknown.
  
  
  RESULTS
  Six cellulolytic enzymes critical for the hydrolysis of crystalline cellulose were purified from T. cellulolyticus culture supernatant using an enzyme assay based on synergistic hydrolysis of Avicel. The purified enzymes were identified by their substrate specificities and analyses of trypsin-digested peptide fragments and were classified into the following glycosyl hydrolase (GH) families: GH3 (β-glucosidase, Bgl3A), GH5 (endoglucanase, Cel5A), GH6 (cellobiohydrolase II, Cel6A), GH7 (cellobiohydrolase I and endoglucanase, Cel7A and Cel7B, respectively), and GH10 (xylanase, Xyl10A). Hydrolysis of dilute acid-pretreated corn stover (PCS) with mixtures of the purified enzymes showed that Cel5A, Cel7B, and Xyl10A each had synergistic effects with a mixture of Cel6A and Cel7A. Cel5A seemed to be more effective in the synergistic hydrolysis of the PCS than Cel7B. The ratio of Cel5A, Cel6A, Cel7A, and Xyl10A was statistically optimized for the hydrolysis of PCS glucan in the presence of Bgl3A. The resultant mixture achieved higher PCS glucan hydrolysis at lower enzyme loading than a culture filtrate from T. cellulolyticus or a commercial enzyme preparation, demonstrating that the five enzymes play a role as core enzymes in the hydrolysis of PCS glucan.
  
  
  CONCLUSIONS
  Core cellulolytic enzymes in the T. cellulolyticus cellulase system were identified to Cel5A, Cel6A, Cel7A, Xyl10A, and Bgl3A and characterized. The optimized mixture of these five enzymes was highly effective for the hydrolysis of PCS glucan, providing a foundation for future improvement of the T. cellulolyticus cellulase system.
authors.affiliation: Biomass Refinery Research Center, National Institute of Advanced Industrial Science and Technology, 3-11-32 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-0046 Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Inoue
authors.affiliation: Biosciences Center, National Renewable Energy Laboratory, 1617 Cole Boulevard, Golden, CO 80401 USA.
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Decker
authors.affiliation: Biosciences Center, National Renewable Energy Laboratory, 1617 Cole Boulevard, Golden, CO 80401 USA.
authors.forename: Larry E
authors.initials: LE
authors.lastname: Taylor
authors.affiliation: Biomass Refinery Research Center, National Institute of Advanced Industrial Science and Technology, 3-11-32 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-0046 Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Yano
authors.affiliation: Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto, 606-8502 Japan.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Sawayama
keywords: Cellulases; Enzymatic hydrolysis; Lignocellulose; Pretreated corn stover; Talaromyces cellulolyticus