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Biochemical Characterization of the Highly Thermostable β-Xylosidase from Caldicellulosiruptor saccharolyticus

Wellalage Don, Dilan Karunathilaka
http://orcid.org/0000-0003-3372-0349
http://rave.ohiolink.edu/etdc/view?acc_num=dayton1565064459365423

Abstract Details

2019, Master of Science (M.S.), University of Dayton, Chemical Engineering.
There has recently been an increasing focus on the conversion of lignocellulosic biomass to biofuel as an alternative to petroleum. The current bottleneck for the process is efficient hydrolysis of lignocellulose into simple sugar molecules for fermentation to ethanol. Xylan represents the major hemicellulose in plants and is identified as the second most abundant polysaccharide on earth. The complete degradation of xylan requires several enzymes working synergistically, including endoxylanases and β-xylosidases. β-xylosidases are capable of hydrolyzing xylo-oligosaccharides to xylose. Thermostable β-xylosidases are more desirable in biofuel production due to their ability to withstand harsh process conditions. This research characterizes glycoside hydrolase enzymes from the extreme thermophilic bacterium Caldicellulosiruptor saccharolyticus, which are predicted to possess the ability to degrade xylan into the fermentable sugar xylose. Thermostable β-xylosidase encoded by Csac_2409 of GH39 from C. saccharolyticus was recombinantly expressed by GenScript and the protein purified to 75% purity. The protein was then characterized to determine the substrate preference, optimal temperature, pH value, thermal stability, and kinetic constants. Thermostable β-xylosidase showed activity over wide range of pH and temperature with optimal pH of 6.5 and temperature of 80°C. The enzyme indicated high thermal stability at 70°C with half-life close to 3 hours. Michaelis-Menten kinetic parameters, KM, V, kcat, and kcat/Km were determined to be 0.918 mM, 0.251 mM/min, 13.6 s-1, and 14.75 s-1mM-1, respectively. Understanding the function and optimal conditions of the enzyme could help the advancement of the lignocellulosic ethanol process, which would ultimately lead to less fossil fuel usage and more environmentally friendly transportation fuels.
Donald Comfort, PhD (Advisor)
76 p.
Chemical Engineering
Biofuel; Lignocellulose; Thermostable; Xylosidase

Recommended Citations

Citations

  • Wellalage Don, D. K. (2019). Biochemical Characterization of the Highly Thermostable β-Xylosidase from Caldicellulosiruptor saccharolyticus [Master's thesis, University of Dayton]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=dayton1565064459365423

    APA Style (7th edition)

  • Wellalage Don, Dilan Karunathilaka. Biochemical Characterization of the Highly Thermostable β-Xylosidase from Caldicellulosiruptor saccharolyticus. 2019. University of Dayton, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=dayton1565064459365423.

    MLA Style (8th edition)

  • Wellalage Don, Dilan Karunathilaka. "Biochemical Characterization of the Highly Thermostable β-Xylosidase from Caldicellulosiruptor saccharolyticus." Master's thesis, University of Dayton, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=dayton1565064459365423

    Chicago Manual of Style (17th edition)

dayton1565064459365423
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This open access ETD is published by University of Dayton and OhioLINK.