Donaghy J, McKay A M
Food Science Division (Microbiology), Department of Agriculture for Northern Ireland, Belfast, UK.
J Appl Microbiol. 1997 Dec;83(6):718-26. doi: 10.1046/j.1365-2672.1997.00307.x.
An extracellular phenolic acid esterase produced by the fungus Penicillium expansum in solid state culture released ferulic and rho-coumaric acid from methyl esters of the acids, and from the phenolic-carbohydrate esters O-[5-O-(trans-feruloyl)-alpha-L-arabinofuranosyl]-(1-->3)-O-beta- D-xylopyranosyl-(1-->4)-D-xylopyranose (FAXX) and O-[5-O-((E)-rho-coumaroyl)-alpha-L-arabinofuranosyl]- (1-->3)-O-beta-D-xylopyranosyl-(1-->4)-D-xylopyranose (PAXX). The esterase was purified 360-fold in successive steps involving ultrafiltration and column chromatography by gel filtration, anion exchange and hydrophobic interaction. These chromatographic methods separated the phenolic acid esterase from alpha-L-arabinofuranosidase, pectate and pectin lyase, polygalacturonase, xylanase and beta-D-xylosidase activities. The phenolic acid esterase had an apparent mass of 65 kDa under non-denaturing conditions and a mass of 57.5 kDa under denaturing conditions. Optimal pH and temperature were 5.6 and 37 degrees C, respectively and the metal ions Cu2+ and Fe3+ at concentrations of 5 mmol 1-1 inhibited feruloyl esterase activity by 95% and 44%, respectively, at the optimum pH and temperature. The apparent Km and Vmax of the purified feruloyl esterase for methyl ferulate at pH 5.6 and 37 degrees C were 2.6 mmol 1-1 and 27.1 mumol min-1 mg-1. The corresponding constants of rho-coumaroyl esterase for methyl coumarate were 2.9 mmol 1-1 and 18.6 mumol min-1 mg-1.
扩展青霉在固态培养中产生的一种胞外酚酸酯酶可从阿魏酸和对香豆酸的甲酯以及酚类 - 碳水化合物酯O - [5 - O - (反式 - 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 (FAXX) 和O - [5 - O - ((E) - 对香豆酰基) - 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 (PAXX) 中释放出阿魏酸和对香豆酸。该酯酶通过超滤和凝胶过滤、阴离子交换及疏水相互作用柱色谱等连续步骤纯化了360倍。这些色谱方法将酚酸酯酶与α - L - 阿拉伯呋喃糖苷酶、果胶酸和果胶裂解酶、多聚半乳糖醛酸酶、木聚糖酶及β - D - 木糖苷酶活性分离开来。该酚酸酯酶在非变性条件下的表观质量为65 kDa，在变性条件下为57.5 kDa。最适pH和温度分别为5.6和37℃，在最适pH和温度下，浓度为5 mmol·L⁻¹的金属离子Cu²⁺和Fe³⁺分别使阿魏酰酯酶活性抑制95%和44%。纯化后的阿魏酰酯酶在pH 5.6和37℃时对阿魏酸甲酯的表观Km和Vmax分别为2.6 mmol·L⁻¹和27.1 μmol·min⁻¹·mg⁻¹。对香豆酰酯酶对香豆酸甲酯的相应常数分别为2.9 mmol·L⁻¹和18.6 μmol·min⁻¹·mg⁻¹。
