朱黄青霉α-1,6-葡聚糖酶在毕赤酵母中的分泌表达

Expression of α-1,6-dextranase from Penicillium minioluteum in P.pastoris

α-1,6-葡聚糖酶是专一作用于α-1,6糖苷键产生小分子葡聚糖的一类水解酶,广泛的运用于制糖工业和啤酒工业中。采用PCR法扩增朱黄青霉(Penicillium minioluteum)C12114的α-1,6-葡聚糖酶基因,将其插入毕赤酵母表达载体pPIC9K。经SacI酶线性化电击转入毕赤酵母基因组,构建重组酵母GS115/pPIC9K-dex。对构建成功的转化子进行1.5%的甲醇诱导表达,在30℃条件下培养7d时酶活达到最大值,为88.35U/mL。

α-1,6-dextranase,which can hydrolyze dextran specifically by cutting off the α-1,6-glycosidic bond to release shorter saccharides,was widely used in many fields such as sugar industry and beer industry.The gene of α-1,6-dextranase（dex）was amplified through PCR by using Penicillium minioluteum C12114 genomic DNA as template.The amplified gene was cloned into vector pPIC9K and the recombinant plasmid pPIC9K-dex was linearzed with Sac I,then transformed into P.pastoris GS115 by electroporation.The positive transformant was induced to express the enzyme with 1.5% methanol for 7 days under the 30℃,and the activity of the enzyme could reach 88.35U/mL.