ARO10基因在Saccharomyces cerevisiae中的克隆表达及其对3-甲硫基丙醇合成代谢的影响

Cloning and Expression of ARO10 Gene in Saccharomyces cerevisiae and It's Influence on 3-(methylthio)-1-propanol Biosynthetic Metabolism

研究了脱羧酶ARO10基因克隆与过量表达对酿酒酵母INVSc1 3-甲硫基丙醇合成途径的代谢流量影响。将脱羧酶基因ARO10与穿梭质粒pYES2连接,构建其酿酒酵母表达质粒(载体)pYES2-ARO10,LiAc/SSD-NA/PEG方法转化酿酒酵母菌株INVSc1中进行表达,验证ARO10基因过量表达对发酵产物3-甲硫基丙醇影响。结果表明,构建的酿酒酵母转化子SC10-1发酵120 h时,3-甲硫基丙醇生成量达到0.90 g/L,与未导入脱羧酶ARO10基因的对照菌株相比,3-甲硫基丙醇产量提高55.2%。因此,S.cerevisiae s288c中脱羧酶(EC 4.1.1.72)是3-甲硫基丙醇生物合成途径的关键限速酶,其增强脱羧酶基因ARO10的克隆及基因表达,有利于提高3-甲硫基丙醇的合成代谢流量。

In this study, the influence of the decarboxylase （EC 4.1.1.72） gene AROIO cloning and overex- pression on the metabolic flux analysis of 3-（methylthio）-1-propanol biosynthesis in Saccharomyces cerevisiae INVScl was investigated. Results showed that the introduction of the recombinant plasmid pYES2-AR010 containing AROIO gene into S. cerevisiae INVScl by using the lithium acetate method resulted in the overexpression of decarboxylase. And the level of 3-（methylthio）-1-propanol biosynthesis in the recombinant strain SC10-1 was significantly enhanced. The maximum yield of 0.90 g/L was achieved when the recombinant strain was incubated for 120 h. Compared with the control strain, the yield increased by 55.2%. These results clearly indicated the decarboxylase from S. cerevisiae s288c is a key rate-limiting enzyme in the biosynthesis of 3-（ methylthio）-l-propanol, and the overexpression of AROIO gene is helpful to the 3-（methylthio）-1-propanol biosynthesis.