Purification and properties of a feruloyl esterase involved in lignocellulose degradation by Aureobasidium pullulans
Date
2003
Authors
Rumbold K.
Biely P.
Mastihubova M.
Gudelj M.
Gubitz G.
Robra K.-H.
Prior B.A.
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Abstract
The lignocellulolytic fungus Aureobasidium pullulans NRRL Y 2311-1 produces feruloyl esterase activity when grown on birchwood xylan. Feruloyl esterase was purified from culture supernatant by ultrafiltration and anion-exchange, hydrophobic interaction, and gel filtration chromatography. The pure enzyme is a monomer with an estimated molecular mass of 210 kDa in both native and denatured forms and has an apparent degree of glycosylation of 48%. The enzyme has a pI of 6.5, and maximum activity is observed at pH 6.7 and 60°C. Specific activities for methyl ferulate, methyl p-coumarate, methyl sinapate, and methyl caffeate are 21.6, 35.3, 12.9, and 30.4 μmol/min/mg, respectively. The pure feruloyl esterase transforms both 2-O and 5-O arabinofuranosidase-linked ferulate equally well and also shows high activity on the substrates 4-O-trans-feruloyl-xylopyranoside, O-{5-O-[(E)-feruloyl]-α-L-arabinofuranosyl}-(1,3)-O-β -D-xylopyranosyl-(1,4)-D-xylopyranose, and p-nitrophenyl-acetate but reveals only low activity on p-nitrophenyl-butyrate. The catalytic efficiency (k cat/Km) of the enzyme was highest on methyl p-coumarate of all the substrates tested. Sequencing revealed the following eight N-terminal amino acids: AVYTLDGD.
Description
Keywords
Amino acids, Catalyst activity, Fungi, Liquid chromatography, Microbiology, Monomers, Plant cell culture, Ultrafiltration, Hydrophobic interaction, Enzymes, 4 nitrophenyl acetate, butyric acid 4 nitrophenyl ester, caffeic acid methyl ester, esterase, ferulic acid methyl ester, feruloyl esterase, hydroxyacid, lignocellulose, methyl 4 coumaric acid, sinapic acid methyl ester, unclassified drug, enzyme activity, fungus, amino terminal sequence, anion exchange, article, Aureobasidium pullulans, enzyme isolation, enzyme purification, fungal metabolism, gel filtration chromatography, microbial degradation, nonhuman, pH, protein glycosylation, species difference, thermostability, ultrafiltration, Ascomycota, Carboxylic Ester Hydrolases, Cellulose, Chromatography, Ion Exchange, Fermentation, Kinetics, Lignin, Substrate Specificity, Ultrafiltration, Aureobasidium, Aureobasidium pullulans, Felis catus, Fungi
Citation
Applied and Environmental Microbiology
69
9
69
9