Mycobacterium sp.LY-1高效转化植物甾醇两相体系的构建  

Constructing the two-phase system for the highly efficient transformation of phytosterols by Mycobacterium sp.LY-1

作　　者：许桠楠 李会 周龙飞 张杰[2] 陈立营 史劲松 许正宏[4] XU Yanan;LI Hui;ZHOU Longfei;ZHANG Jie;CHEN Liying;SHI Jinsong;XU Zhenghong(School of Pharmaceutical Science,Jiangnan University,Wuxi 214122,China;Tianjin Tianyao Pharmaceuticals Co.,Ltd.,Tianjin 300462,China;Tianjin Pharmaceutical Research Institute Co.,Ltd.,Tianjin 300462,China;Key Laboratory of Industrial Biotechnology of Ministry of Education,School of Bioengineering,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学药学院,无锡214122 [2]天津天药药业股份有限公司,天津300462 [3]天津药业研究院股份有限公司,天津300462 [4]工业生物技术教育部重点实验室,江南大学生物工程学院,无锡214122

出　　处：《应用与环境生物学报》2020年第4期747-752,共6页Chinese Journal of Applied and Environmental Biology

基　　金：国家重点研发计划项目(2019YFA0905300);天津市合成生物技术创新能力提升行动项目(TSBICIP-KJGG-001-14);中国博士后科学基金面上项目(2018M640452);江苏省研究生科研与实践创新计划项目(SJCX18_0640)资助。

摘　　要：9α-羟基雄甾-4-烯-3,17-二酮(9α-OH-AD)是一种卤代皮质类激素的重要前体,具有重要的商用价值.为进一步提高目的产物9α-OH-AD的产量,在本实验室前期利用非离子表面活性剂壬基酚乙氧乙烯醚(TX-40)替代大豆油进行Mycobacterium sp.LY-1转化植物甾醇的基础上,通过考察16种助溶剂与TX-40复配对菌株LY-1转化植物甾醇生产9α-OH-AD的影响,确定最适助溶剂为司盘-20.通过正交优化实验确定司盘-20的最优添加浓度为0.1%.进一步考察不同浓度细胞膜结构抑制剂甘氨酸、鱼精蛋白、异烟肼对菌株LY-1转化植物甾醇的影响,结果显示在以上转化体系中添加1.0μg/mL的鱼精蛋白时,9α-OH-AD的摩尔得率最高,达到46.4%.最终确定菌株LY-1降解植物甾醇的两相转化体系为TX-400.7%,司盘-200.1%,鱼精蛋白1.0μg/mL.该体系于植物甾醇的投料浓度提高至35 g/L时,目的产物9α-OH-AD的摩尔得率仍能稳定在32.5%以上.本研究成功构建了Mycobacterium sp.LY-1高效转化植物甾醇的两相体系,结果可为其工业化应用奠定良好基础.(图4表2参26)The 9α-hydroxyandrost-4-ene-3,17-dione(9α-OH-AD)is an important precursor of halogenated corticosteroids and has an important commercial value.The non-ionic surfactant nonylphenol ethoxyethylene ether(TX-40)was investigated in our previous studies and used in phytosterols transformation instead of soybean oil by Mycobacterium sp.LY-1.To further increase the yield of the target product 9α-OH-AD,the effects of the complex formulation of the 16 co-solvent and the TX-40 on phytosterol transformation to 9α-OH-AD by the LY-1 strain were studied,where the most suitable compound was screened as Span-20.The optimal concentration of Span-20 was determined to be 0.1%as shown by the orthogonal optimization experiment.Hence,based on this work,the effects of the different concentrations of the cellular membrane structure inhibitors,such as glycine,protamine,and isoniazid on the transformation of phytosterols by the LY-1 strain were also investigated.Our results showed that the highest molar yield of 9α-OH-AD was 46.4%when 1.0μg/mL of protamine was added to the aforementioned transformation system.Finally,we determined that the two-phase transformation system for the degradation of phytosterol by the LY-1 strain was when TX-40 was 0.7%,Span-20 was 0.1%,and protamine was 1.0μg/mL.In addition,when the concentration of phytosterol was increased to 35 g/L,the molar yield of the target product 9α-OH-AD was still stable at 32.5%,which sets a promising foundation for the industrial application of this system.

关 键 词：Mycobacterium sp.LY-1 两相转化体系 9α-羟基雄甾-4-烯-3 17-二酮 细胞膜结构抑制剂 正交优化 

分 类 号：TQ460.1[化学工程—制药化工]