Department of Applied Microbiology, Lund University, P.O. Box 124, SE-22100 Lund, Sweden.
Microb Cell Fact. 2010 May 27;9:40. doi: 10.1186/1475-2859-9-40.
In Saccharomyces cerevisiae galactose is initially metabolized through the Leloir pathway after which glucose 6-phosphate enters glycolysis. Galactose is controlled both by glucose repression and by galactose induction. The gene PGM2 encodes the last enzyme of the Leloir pathway, phosphoglucomutase 2 (Pgm2p), which catalyses the reversible conversion of glucose 1-phosphate to glucose 6-phosphate. Overexpression of PGM2 has previously been shown to enhance aerobic growth of S. cerevisiae in galactose medium.
In the present study we show that overexpression of PGM2 under control of the HXT7'promoter from an integrative plasmid increased the PGM activity 5 to 6 times, which significantly reduced the lag phase of glucose-pregrown cells in an anaerobic galactose culture. PGM2 overexpression also increased the anaerobic specific growth rate whereas ethanol production was less influenced. When PGM2 was overexpressed from a multicopy plasmid instead, the PGM activity increased almost 32 times. However, this increase of PGM activity did not further improve aerobic galactose fermentation as compared to the strain carrying PGM2 on the integrative plasmid.
PGM2 overexpression in S. cerevisiae from an integrative plasmid is sufficient to reduce the lag phase and to enhance the growth rate in anaerobic galactose fermentation, which results in an overall decrease in fermentation duration. This observation is of particular importance for the future development of stable industrial strains with enhanced PGM activity.
在酿酒酵母中,半乳糖最初通过 Leloir 途径代谢,然后葡萄糖 6-磷酸进入糖酵解。半乳糖受到葡萄糖抑制和半乳糖诱导的双重控制。PGM2 基因编码 Leloir 途径的最后一种酶,磷酸葡萄糖变位酶 2(Pgm2p),它催化葡萄糖 1-磷酸可逆转化为葡萄糖 6-磷酸。先前的研究表明,PGM2 的过表达可增强酿酒酵母在半乳糖培养基中的好氧生长。
本研究表明,在整合质粒的 HXT7'启动子控制下过表达 PGM2,可使 PGM 活性提高 5 至 6 倍,这显著缩短了葡萄糖预培养细胞在无氧半乳糖培养中的迟滞期。PGM2 的过表达还提高了厌氧比生长速率,而乙醇的产量则受影响较小。当 PGM2 从多拷贝质粒过表达时,PGM 活性增加了近 32 倍。然而,与携带整合质粒上 PGM2 的菌株相比,这种 PGM 活性的增加并没有进一步改善好氧半乳糖发酵。
在酿酒酵母中,整合质粒上的 PGM2 过表达足以缩短迟滞期并增强厌氧半乳糖发酵中的生长速率,从而整体缩短发酵时间。这一观察结果对于未来开发具有增强 PGM 活性的稳定工业菌株具有重要意义。