Purification and properties of the xylanase produced by Thermomyces lanuginosus
Purification and properties of the xylanase produced by Thermomyces lanuginosus
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DOI：
10.1016/0141-0229(95)00248-0
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发表时间：
1996-09-01
影响因子：
3.4
通讯作者：
Mrsa, V
中科院分区：
文献类型：
--
作者：
Cesar, T;Mrsa, V
对德国典型菌种中的一株编号为DSM 5826的发酵菌--羊毛热霉菌深层发酵所得的木聚糖酶进行了层析纯化，并对其进行了性质鉴定。分别用25.5 kDa、24.0 kDa和22.5 kDa的SDS-PAGE、梯度凝胶电泳法和凝胶过滤法测定了该酶的相对分子质量。测得该酶的等电点为pH 4.1。结果表明，该酶以内切木聚糖酶的形式对木聚糖进行水解性研究，几乎不具有纤维分解或其他类似的水解酶活性。在60-70℃的温度范围内，该酶在pH为7.0左右时表现出最高的活力。由于二硫双-2-硝基苯甲酸、对羟基汞苯甲酸和Hg2+离子对该酶有完全的抑制作用，因此该酶只含有一个对木聚糖酶活性重要的半胱氨酸残基。Mn2+、Fe2+和β-巯基乙醇对木聚糖酶活性有促进作用，其中Fe2+和β-巯基乙醇联合作用效果最好。木聚糖酶在pH为5.0-9.0，温度高达60℃时稳定96h，添加甘油、β-巯基乙醇或聚乙二醇能显著稳定木聚糖酶。
Xylanase obtained by the submersed fermentation of Thermomyces lanuginosus, a strain classified in the German type culture collection under the number DSM 5826, was purified to homogeneity by a combination of chromatographic methods and characterized. The molecular mass of the enzyme was estimated by SDS-electrophoresis, gradient gel electrophoresis, or gel filtration at 25.5 kDa, 24.0 kDa, or 22.5 KDa, respectively. The isoelectric point of the enzyme was determined to be pH 4.1. It was found that the enzyme hydrolyzed xylan as an endoxylanase and had practically no cellulolytic or any other similar hydrolytic activity. It exhibited the highest activity at a pH around 7.0 and in the temperature range of 60-70 degrees C, It was found that the enzyme contains only one cysteine residue which was important for the xylanase activity, since dithiobis-2-nitrobenzoic acid, p-hydroxymercuribenzoic acid, and Hg2+ ions completely inhibited the enzyme. Mn2+, Fe2+, and beta-mercaptoethanol enhanced the xylanase activity; the best results were obtained by the combined action of Fe2+ ions and beta-mercaptoethanol. Xylanase was stable for 96 h at a pH between 5.0-9.0 and at temperatures up to 60 degrees C. Significant stabilization of the enzyme was achieved by the addition of glycerol, beta-mercaptoethanol, or polyethylene glycol.