Abstract
The gene encoding a glycoside hydrolase family 43 enzyme termed deAX was isolated and subcloned from a culture seeded with a compost starter mixed bacterium population, expressed with a C-terminal His6-tag, and purified to apparent homogeneity. deAX was monomeric in solution and had a broad pH maximum between pH 5.5 and pH 7. A twofold greater k cat/K m for the p-nitrophenyl derivative of α-l-arabinofuranose versus that for the isomeric substrate β-d-xylopyranose was due to an appreciably lower K m for the arabinofuranosyl substrate. Substrate inhibition was observed for both 4-methylumbelliferryl arabinofuranoside and the xylopyranoside cogener. While no loss of activity was observed over 4 h at 40°C, the observed t 1/2 value rapidly decreased from 630 min at 49°C to 47 min at 53°C. The enzyme exhibited end-product inhibition, with a K i for xylose of 145 mM, 18.5 mM for arabinose, and 750 mM for glucose. Regarding natural substrate specificity, deAX had arabinofuranosidase activity on sugar beet arabinan, 1,5-α-l-arabinobiose, and 1,5-α-l-arabinotriose, and wheat and rye arabinoxylan, while xylosidase activity was detected for the substrates xylobiose, xylotriose, xylotetraose, and arabinoxylan from beech and birch. Thus, deAX can be classified as a dual-function xylosidase/arabinofuranosidase with respect to both artificial and natural substrate specificity.
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The authors would like to thank Gregory Gray for GLC analysis. Reference to a company and/or products is for purposes of information and does not imply approval or recommendation of the product to the exclusion of others which may also be suitable. All programs and services of the US Department of Agriculture are offered on a non-discriminatory basis without regard to race, color, national origin, religion, sex, age, marital status, or handicap.
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Wagschal, K., Heng, C., Lee, C.C. et al. Biochemical characterization of a novel dual-function arabinofuranosidase/xylosidase isolated from a compost starter mixture. Appl Microbiol Biotechnol 81, 855–863 (2009). https://doi.org/10.1007/s00253-008-1662-4
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DOI: https://doi.org/10.1007/s00253-008-1662-4