Title: Differentiation of feruloyl esterases on synthetic substrates in alpha-arabinofuranosidase-coupled and ultraviolet-spectrophotometric assays Biely P, Mastihubova M, van Zyl WH, Prior BA Ref: Analytical Biochemistry, 311:68, 2002 : PubMed
4-Nitrophenyl 5-O-trans-feruloyl-alpha-L-arabinofuranoside and 4-nitrophenyl 2-O-trans-feruloyl-alpha-L-arabinofuranoside, synthesized by our group (M. Mastihubov, J. Szemesov, and P. Biely), were found to be suitable substrates for determination of activity of feruloyl esterases (FeEs) exhibiting affinity for 5-O- and 2-O-feruloylated alpha-L-arabinofuranosyl residues. One assay is based on coupling the FeE-catalyzed formation of 4-nitrophenyl alpha-L-arabinofuranoside with its efficient hydrolysis by alpha-L-arabinofuranosidase to release 4-nitrophenol. An alternative assay explores the difference in the molar absorbances at 340 nm of the substrate (ferulic acid esters) and the reaction products, which are (1) free ferulic acid and 4-nitrophenyl alpha-L-arabinofuranoside in samples free of alpha-L-arabinofuranosidase and (2) ferulic acid, 4-nitrophenyl alpha-L-arabinofuranoside, and/or 4-nitrophenol in samples containing alpha-L-arabinofuranosidase. The new substrates represent convenient tools to differentiate FeEs on the basis of substrate specificity.
Biely P, Mastihubova M, van Zyl WH, Prior BA (2002) Differentiation of feruloyl esterases on synthetic substrates in alpha-arabinofuranosidase-coupled and ultraviolet-spectrophotometric assays Analytical Biochemistry311: 68-75
Biely P, Mastihubova M, van Zyl WH, Prior BA (2002) Analytical Biochemistry311: 68-75
