毛云芝菌漆酶的分离纯化和性质研究

Purification and Properties of Laccase Produced by Coriolus hirsutus

将发酵液经真空抽滤后得粗酶液,以硫酸铵为盐析剂进行分级盐析沉淀、再经DEAE-52离子交换层析和Sephadex G-75凝胶过滤层析对漆酶进行分离纯化,用SDS-PAGE证明可获得纯的漆酶,纯化倍数为183.69倍,酶活回收率为24.77%。实验表明,该酶在30℃条件下具有较高的稳定性;在pH3.4～5.8范围内的相对酶活均在80%以上,漆酶稳定性较好,pH3.8的时候漆酶的稳定性最好。

Crude laccase was collected after fermentation broth was filtrated with vacuum pump, and then purified to electrophoretic homogeneity by the steps of ammonium sulfate precipitation, ion-exchange chromatography on DEAE-cellulose and gel filtration on Sephadex G-75. The purification of about 183.69 folds is achieved with an overall yield of 24.77%. Its molecular weight is estimated to be about 60.3 kD by SDS-PAGE. Studies on the properties of the laccase showed that enzyme activity is more stable when preserved at 30 ℃. The relative activity of laccase is above 80% when preserved at the pH range of 3.4 to 5.8, and its stability is the highest when preserved at the pH of 3.8.