State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China.
Carbohydr Polym. 2014 Jan 30;101:89-95. doi: 10.1016/j.carbpol.2013.09.011. Epub 2013 Sep 12.
Protopectinase is an enzyme that solubilizes protopectin forming highly polymerized soluble pectin. Protopectinase activity was detected from Aspergillus oryzae PO isolated from soil of persimmon orchard. Response surface methodology of Box-Behnken Design with three fermentation variables (temperature, NaNO3 and apple pomace concentration) was used to optimize protopectinase production of A. oryzae PO, and protopectinase activity was improved to 270.0 U/ml. Endo-polygalacturonase belonged to A-type PPase from A. oryzae PO was cloned and expressed in Pichia pastoris GS115. The endo-polygalacturonase expression was 0.418 mg/ml and the specific activity of purified recombinant endo-polygalacturonase was 7520 U/mg toward polygalacturonic acid. The optimal temperature and pH of recombinant endo-polygalacturonase were 45°C and 5.0, respectively. The recombinant endo-polygalacturonase activity was enhanced by the presence of Mg(2+), while Ca(2+), Ni(2+) Mn(2+), Cu(2+) and SDS strongly inhibited the enzyme activity. The apparent Km value and Vmax value were 5.59 mg/ml and 1.01 μmol/(minml), respectively.
原果胶酶是一种能够溶解原果胶并形成高度聚合的可溶性果胶的酶。原果胶酶活性是从柿子园土壤中分离的米曲霉 PO 中检测到的。采用 Box-Behnken 设计的响应面法，对温度、NaNO3 和苹果渣浓度三个发酵变量进行优化，使米曲霉 PO 的原果胶酶活性提高到 270.0 U/ml。内切聚半乳糖醛酸酶属于 A 型 PPase，从米曲霉 PO 中克隆并在毕赤酵母 GS115 中表达。内切聚半乳糖醛酸酶的表达量为 0.418 mg/ml，纯化的重组内切聚半乳糖醛酸酶对聚半乳糖醛酸的比活性为 7520 U/mg。重组内切聚半乳糖醛酸酶的最适温度和 pH 分别为 45°C 和 5.0。Mg2+存在时可增强重组内切聚半乳糖醛酸酶的活性，而 Ca2+、Ni2+、Mn2+、Cu2+和 SDS 强烈抑制酶活性。该酶的表观 Km 值和 Vmax 值分别为 5.59 mg/ml 和 1.01 μmol/(minml)。
