A screening method for endo-beta-1,4-xylanase substrate selectivity.
Endoxylanase (EC 3.2.1.8) substrate selectivity, i.e., its relative activity toward water-unextractable arabinoxylan (WU-AX) and water-extractable arabinoxylan (WE-AX) substrates, is important for its functionality in biotechnological processes such as bread-making and gluten starch separation. A screening method for rapidly determining said substrate selectivity was developed. Endoxylanase activity toward WU-AX was estimated by incubation of insoluble chromogenic substrate with a range of enzyme concentrations in microtiter plates, followed by colorimetric measurement of the dye released in the supernatant. A similar approach using soluble substrate and ethanol precipitation of unhydrolysed AX fragments was used to estimate enzyme activity toward WE-AX. A substrate selectivity factor was defined as the ratio of enzyme activity toward insoluble substrate over enzyme activity toward soluble substrate. A Bacillus subtilis and an Aspergillus aculeatus endoxylanase, known to have widely varying relative rates of hydrolysis of WU-AX and WE-AX, varied most in their substrate selectivity, while the endoxylanases of Aspergillus niger, Trichoderma longibrachiatum, and Trichoderma viride displayed intermediate such relative activities.
Moers
Karolien
K
Laboratory of Food Chemistry, Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium. karolien.moers@agr.kuleuven.ac.be
Courtin
Christophe M
CM
Brijs
Kristof
K
Delcour
Jan A
JA
D001230:Aspergillus / Q000201:enzymology; D001412:Bacillus subtilis / Q000201:enzymology; D043364:Endo-1,4-beta Xylanases / Q000378:metabolism*; D007700:Kinetics; D013054:Spectrophotometry, Atomic; D013379:Substrate Specificity; D014242:Trichoderma / Q000201:enzymology; D014990:Xylans / Q000378:metabolism*
D014990:Xylans; D043364:Endo-1,4-beta Xylanases