Journal
APPLIED AND ENVIRONMENTAL MICROBIOLOGY
Volume 79, Issue 5, Pages 1481-1490Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/AEM.03265-12
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- Energy Biosciences Institute (EBI)
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Xylose, the major constituent of xylans, as well as the side chain sugars, such as arabinose, can be metabolized by engineered yeasts into ethanol. Therefore, xylan-degrading enzymes that efficiently hydrolyze xylans will add value to cellulases used in hydrolysis of plant cell wall polysaccharides for conversion to biofuels. Heterogeneous xylan is a complex substrate, and it requires multiple enzymes to release its constituent sugars. However, the components of xylan-degrading enzymes are often individually characterized, leading to a dearth of research that analyzes synergistic actions of the components of xylan-degrading enzymes. In the present report, six genes predicted to encode components of the xylan-degrading enzymes of the thermophilic bacterium Caldicellulosiruptor bescii were expressed in Escherichia coli, and the recombinant proteins were investigated as individual enzymes and also as a xylan-degrading enzyme cocktail. Most of the component enzymes of the xylan-degrading enzyme mixture had similar optimal pH (5.5 to similar to 6.5) and temperature (75 to similar to 90 degrees C), and this facilitated their investigation as an enzyme cocktail for deconstruction of xylans. The core enzymes (two endoxylanases and a beta-xylosidase) exhibited high turnover numbers during catalysis, with the two endoxylanases yielding estimated k(cat) values of similar to 8,000 and similar to 4,500 s(-1), respectively, on soluble wheat arabinoxylan. Addition of side chain-cleaving enzymes to the core enzymes increased depolymerization of a more complex model substrate, oat spelt xylan. The C. bescii xylan-degrading enzyme mixture effectively hydrolyzes xylan at 65 to 80 degrees C and can serve as a basal mixture for deconstruction of xylans in bioenergy feedstock at high temperatures.
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