Ochrobacterum intermedium DN2新型烟碱降解酶的分离纯化

Purification of A Novel Nicotine-degradation Enzyme in Ochrobacterum intermedium DN2

中间苍白杆菌(OchrobacterumintermediumDN2)是一株新发现的可以降解烟碱的菌株。本研究对O.intermediumDN2烟碱降解酶的合成动态进行了分析,确定其酶的合成模式为同步合成型。另外,对它的烟碱降解酶进行了分离纯化。粗酶经硫酸铵沉淀、SephadexG-100分子筛层析、DEAE-SephadexA50离子交换层析等步骤后获得电泳纯的烟碱降解酶,纯化倍数35.24倍,酶活回收率6.4%。经SDS-PAGE电泳确定为电泳纯,测得该酶的分子量约为40.06kDa。由中间苍白杆菌产生的烟碱降解酶的分离纯化在国内外还是首次报道。

In previous study, it was found that Ochrobacterum intermedium DN2 is a kind of bacteria which can utilize nicotine as its sole carbon source. In this work, the production and purification of nicotine-degradation enzyme in O.intermedium DN2 was investigated. The production of nicotine-degradation enzyme was synchronous with cell growth. The enzyme was purified by successive steps of ammonium sulfate precipitation, gel filtration on Sephadex G-100, ion-exchange chromatography on DEAE-Sephadex A50. The enzyme was purified 35.24 folds and was taken back 6.4% of its activity. The purified nicotinedegradation enzyme showed homogeneity on the SDS-PAGE. The molecule weight of this enzyme was determined as being 40.06kDa by SDS-PAGE. This is the fixst report on the production and purification of the nicotine-degradation enzyme in O. intermedium DN2.