一株酸性糖化酶的分离纯化及酶学性质研究

Purification of an Acidic Glucoamylase from Aspergillus niger and its Enzymatic Properties

摘要:

从土壤中筛选得到一株产酸性糖化酶的黑曲霉ASP-S21，粗酶液通过(NH4)2SO4沉淀、Sepharose HP阴离子交换层析、Sephacryl S-200层析柱分离纯化，SDS-PAGE电泳测定其分子量为120 kDa。该酶最适作用温度为65 ℃；酶的最适反应pH值为4.0；在pH 2.2~7.6之间，具有较好的酸稳定性；酶的Km值为0.94 mg/mL，Vmax=142.43 mol(Glu)/min?L。Cu2+和Co2+对酶活有较强的促进作用，10 mM的Cu2+使酶活力提高到129%，Fe3+对酶催化活力抑制作用较强。该酶且具有部分降解生淀粉的能力，在pH 4.0，50 ℃反应 1h，生淀粉酶活力为0.39 U，RDA值为4.57%。得到的黑曲霉ASP-S21酸性糖化酶，产生的糖化酶活力高、耐酸稳定性好，酶学性质符合淀粉糖化工业化过程中对酶的要求，具有良好的研究前景。

Abstract:

An acidic glucoamylase producing strain Aspergillus niger ASP-S21 was isolated from soil. Glucoamylase was purified to homogeneity by ammonium sulfate precipitation ,ion-exchange chromatography with a Sepharose HP column ,and column chromatography with a Sephacryl S-200 column, the molecular weight of SDS-PAGE was 120 kDa. The study of its enzymatic properties showed that the optimum temperature and pH for glucoamylase of such strain were 65 ℃ and 4.0, respectively. Besides, it had good acid stability within the range of pH value from 2.2 to7.6. The Km for glucoamylase was calculated to be 0.94 mg/mL，and the Vmax was 142.43 mol(Glu)/min?L. The enzyme activity was enhanced by Cu2+ and Co2+, and the 10 mM of Cu2+ could improve the glucoamylase activity to 129%. However, the enzyme was strongly inhibited by Fe3+. Its RSGA activities were 0.39U and the RDA values were 4.57%. The glucoamylase was suitable for applying in starch sugar industry.