Kanna et al. AMB Express 2011, 1:15 http://www.amb-express.com/content/1/1/15
Enhancement of b-xylosidase productivity in cellulase producing fungus Acremonium cellulolyticus
Machi Kanna1, Shinichi Yano1*, Hiroyuki Inoue1, Tatsuya Fujii1 and Shigeki Sawayama1,2
Enzymatic hydrolysis is one of the most important processes in bioethanol production from lignocellulosic biomass. Acremonium cellulolyticus is a filamentous fungus with high cellulase production but productivity of hemicellulase, especially b-xylosidase, is lower than other filamentous fungi. We identified 2.4 Kb b-xylosidase gene in the A. cellulolyticus genome sequence information and it encoded 798 amino acids without introns. To enhance hemicellulase productivity in A. cellulolyticus, we transformed this fungus with the identified b-xylosidase gene driven by the cellobiohydrolase Ι (cbh1) promoter, using the protoplast-polyethyleneglycol (PEG) method, and obtained a transformant, YKX1. Hydrolysis rate of xylooligosaccharides was more than 50-fold higher using culture supernatant from YKX1 than that from the parental strain, Y-94. Total cellulase activity (measured by filter paper assay) in YKX1 was not affected by the cbh1 promoter used for expression of b-xylosidase, and induced by cellulose. Since YKX1 can produce larger amount of b-xylosidase without affecting cellulase productivity, it is considered to be beneficial for practical monosaccharide recoveries from lignocellulosic biomass.
Keywords: Hemicellulase, beta-xylosidase, Acremonium cellulolyticus, Transformation, Cellulase
Introduction Ethanol produced from lignocellulosic biomass is a sec- ond-generation biofuel which does not compete with food resources (Sims et al. 2010) and is expected to be an alternative to gasoline that reduces dependence on fossil fuels. Bioethanol can be produced from lignocellu- losic biomass via several processes; pretreatment, enzy- matic hydrolysis, and fermentation. In these processes, effective enzymatic hydrolysis of cellulose and hemicel- lulose is the most important step. Therefore, we focused on the enzyme activities that catalyze the saccharifica- tion of cellulose and hemicellulose.
Cellulose is the primary component of lignocellulosic biomass. After cellulose, hemicellulose is the second most abundant component of the plant cell wall, and accounts for 20-30% of lignocellulosic biomass (Girio et al. 2010,). Glucuronoxylans (O-acetyl-4-O-methylglucuronoxylan)
are the most abundant type of hemicellulose, and they make up 15-30% of the dry mass in hardwoods (Girio et al. 2010). Although conventional ethanol fermenting yeasts cannot utilize xylose, we have developed efficient xylose- fermentable Saccharomyces cerevisiae strains (Matsushika et al., 2009). Hence, effective saccharification of xylan is practically important for attaining higher yields of monosaccharides.
Endo-b-1, 4-xylanase and b-xylosidase catalyze the pro- duction of xylooligosaccharides from xylan and xylose from xylooligosaccharides, respectively. b-xylosidase hydrolyzes the non-reducing end of xylooligosaccharides. Although purified b-xylosidase is commercially available, it is too costly for practical large-scale applications. It will be beneficial if cellulase-producing filamentous fungi also produce hemicellulase for efficient and cost-effective hydrolysis of lignocellulosic biomass.
* Correspondence: firstname.lastname@example.org Biomass Technology Research Center, National Institute of Advanced Industrial Science and Technology, 3-11-32 Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-0046 Japan Full list of author information is available at the end of the article 1
The b-xylosidase genes have been sequenced and characterized from many species of filamentous fungi. xylA of Aspergillus oryzae (Kitamoto et al. 1999) and xyl I of Aureobasidium pullulans strain ATCC 20524 (Ohta et al. 2010) belong to the glycosyl hydrolase (GH)
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