摘要
由本实验室筛选得到的摩尔摩根氏菌J-8菌株可将底物1-苯基-2-甲氨基丙酮专一性地转化为d-伪麻黄碱。以M.morganiiJ-8为出发菌株,菌体超声破碎后,经硫酸铵沉淀、Phenyl Superose疏水柱层析、DEAD阴离子柱层析和非变性凝胶电泳四步纯化获得电泳纯羰基不对称还原酶。亚基分子质量为42.5kD,高效液相色谱分析酶的分子质量约为84.1kD,初步认为该酶为二聚体蛋白。对所得到的部分纯化酶的酶学性质做了初步研究,纯酶进行基质辅助激光解析电离-飞行质谱分析,比对结果显示为与亮氨酸脱氢酶蛋白有很高相似性。
The purification and the characteristics of an enzyme from Morganella morganii J-8, which could produce d-pseudoephedrine from 1-phenyl-2-methylamine-acetone, were performed in this study. In this research, first, cells were disrupted by ultrasonic treatment at 4℃. The carbonyl enantioselective reductase was purified with a combination of ammonium precipitation, Phenyl Superose hydrophobic chromatography, DEAE anion exchange, and native polyacrylamide gel electrophoresis. The molecular mass of the purified enzyme subunit was estimated to be 42.5kD on sodium dodecyl sulfate-polyacrylamide electrophoresis （SDS-PAGE） . The native molecular mass of the enzyme that was analyzed by high-performance liquid chromatography was found out to be 84.1 kD, which indicated that the enzyme was a dimmer. The purified enzyme was analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and the result showed that the purified enzyme had high homology with leucine dehydrogenase.
出处
《生物工程学报》
CAS
CSCD
北大核心
2007年第2期268-272,共5页
Chinese Journal of Biotechnology
基金
江苏省自然科学基金(No.BK2005013)
高等学校博士学科点专项科研基金(No.20040295002)资助~~
