Kirk N, Piper P W
Department of Biochemistry and Molecular Biology, University College London, UK.
Appl Microbiol Biotechnol. 1994 Nov;42(2-3):340-5. doi: 10.1007/BF00902739.
The signal sequences of the MF alpha 1 prepro alpha-factor gene are frequently used to direct secretion of heterologous proteins from Saccharomyces cerevisiae. They are often employed together with the MF alpha 1 promoter in secretion vectors, such that this promoter directs the transcription of many heterologous gene cassettes in yeast. Most of the existing literature indicates that the MF alpha 1 promoter is constitutive in MAT alpha cells, although some data suggests that it may be more active in respiratory or late logarithmic fermentative cultures. To identify whether there is a growth rate or medium control over MF alpha 1 promoter activity a strain was constructed with an integrated MF alpha 1 promoter-beta-galactosidase (lacZ) reporter gene fusion. Intracellular beta-galactosidase of this strain during batch culture on glucose, raffinose and acetate showed that MF alpha 1 promoter activity was higher during respiratory growth on acetate as compared to more rapid fermentative growth on glucose or raffinose, a result that might indicate this activity being inversely related to growth rate. Chemostat culture confirmed that growth rate does indeed influence MF alpha 1 promoter activity in glucose-grown cells, the activity of this promoter increasing 2- to 2.5-fold as dilution (growth) rates were reduced from maximal values to 0.2 h-1, but then decreasing with the further decreases in dilution rate needed for fully respiratory growth. Thus a promoter generally thought to be constitutive in MAT alpha cells is nevertheless subject to a complex growth rate control.
MFα1前原α因子基因的信号序列常被用于指导酿酒酵母中外源蛋白的分泌。它们常与MFα1启动子一起用于分泌载体中，使得该启动子指导酵母中许多异源基因盒的转录。现有大多数文献表明，MFα1启动子在MATα细胞中是组成型的，尽管一些数据表明它在呼吸或对数后期发酵培养中可能更活跃。为了确定MFα1启动子活性是否受生长速率或培养基的控制，构建了一个带有整合的MFα1启动子-β-半乳糖苷酶（lacZ）报告基因融合体的菌株。该菌株在葡萄糖、棉子糖和乙酸盐分批培养过程中的细胞内β-半乳糖苷酶表明，与在葡萄糖或棉子糖上更快的发酵生长相比，在乙酸盐上呼吸生长期间MFα1启动子活性更高，这一结果可能表明该活性与生长速率呈负相关。恒化器培养证实，生长速率确实会影响葡萄糖培养细胞中MFα1启动子的活性，当稀释（生长）速率从最大值降至0.2 h-1时，该启动子的活性增加2至2.5倍，但随后随着完全呼吸生长所需的稀释速率进一步降低而降低。因此，一个通常被认为在MATα细胞中是组成型的启动子，仍然受到复杂的生长速率控制。
