Rumbold Karl, Biely Peter, Mastihubová Maria, Gudelj Marinka, Gübitz Georg, Robra Karl-Heinz, Prior Bernard A
Department of Microbiology, University of Stellenbosch, 7602 Matieland, South Africa.
Appl Environ Microbiol. 2003 Sep;69(9):5622-6. doi: 10.1128/AEM.69.9.5622-5626.2003.
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 degrees C. Specific activities for methyl ferulate, methyl p-coumarate, methyl sinapate, and methyl caffeate are 21.6, 35.3, 12.9, and 30.4 micro mol/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]-alpha-L-arabinofuranosyl]-(1,3)-O-beta-D-xylopyranosyl-(1,4)-D-xylopyranose, and p-nitrophenyl-acetate but reveals only low activity on p-nitrophenyl-butyrate. The catalytic efficiency (k(cat)/K(m)) of the enzyme was highest on methyl p-coumarate of all the substrates tested. Sequencing revealed the following eight N-terminal amino acids: AVYTLDGD.
木纤维素分解真菌出芽短梗霉NRRL Y 2311-1在以桦木木聚糖为培养基生长时会产生阿魏酸酯酶活性。通过超滤、阴离子交换、疏水相互作用和凝胶过滤色谱法从培养上清液中纯化出阿魏酸酯酶。该纯酶在天然和变性形式下均为单体，估计分子量为210 kDa，表观糖基化程度为48%。该酶的pI为6.5，在pH 6.7和60℃时观察到最大活性。对阿魏酸甲酯、对香豆酸甲酯、芥子酸甲酯和咖啡酸甲酯的比活性分别为21.6、35.3、12.9和30.4微摩尔/分钟/毫克。纯阿魏酸酯酶对2-O和5-O阿拉伯呋喃糖苷连接的阿魏酸的转化效果同样良好，并且对4-O-反式-阿魏酰基-木吡喃糖苷、O-[5-O-[(E)-阿魏酰基]-α-L-阿拉伯呋喃糖基]-(1,3)-O-β-D-木吡喃糖基-(1,4)-D-木吡喃糖和对硝基苯基乙酸酯等底物也表现出高活性，但对对硝基苯基丁酸酯仅表现出低活性。在所有测试底物中，该酶对香豆酸甲酯的催化效率（k(cat)/K(m)）最高。测序显示了以下八个N端氨基酸：AVYTLDGD。
