高效转化植物甾醇为9α-OH-AD的分枝杆菌诱变选育及工艺优化

Mutation breeding of high 9α-hydroxy-androst-4-ene-3,17- dione transforming strains from phytosterols and their conversion process optimization

摘要:

以实验室前期筛选得到的分枝杆菌(Mycobacterium sp. LY-1)为出发菌株，采用等离子诱变(ARTP)技术选育出能高效转化植物甾醇为重要甾体药物中间体9α-羟基雄甾-4-烯-3,17-二酮(9α-OH-AD)的菌株，并对其转化工艺进行优化。经过ARTP诱变选育得到遗传稳定性较好的分枝杆菌突变株Mycobacterium sp. C33，当底物植物甾醇投料浓度为15 g/L时，转化生成9α-OH-AD的摩尔得率达到15.5%，较原始菌株提高34.8%。继而采用正交实验设计方法，对突变菌的发酵培养基组分进行优化，并建立了油水双相转化体系，进一步提高了突变菌株C33的产物摩尔得率，最高达到47.0%，较优化前(15.5%)提高了2倍。

Abstract:

In order to improve transformation efficiency of phytosterols into 9α-hydroxylation of 4-androstene-3,17-dione (9α-OH-AD) by Mycobacterium sp. LY-1, we studied the strains breeding using atmospheric and room temperature plasma (ARTP) technology and optimized their conversion process. A high production strain named C33 with a good genetic stability was selected and the product molar yield reached to 15.5%, 34.8% higher than that of original strain with 15 g/L phytosterols. Furthermore, the fermentation medium was optimized through the design of orthogonal experiment. Besides, oil-water bidirectional transformation system was set up to improve the 9α-OH-AD molar yield of mutant strain C33. With adding 12 mL soybean oil to each 1 g phytosterols, the molar yield of 9α-OH-AD reached 47.0%, which increased twice than that of control (15.5%).