Analysis of cloned SUC2 gene expression in continuous culture of recombinant Saccharomyces cerevisiae

✍ Scribed by Jae K. Jang; Yu R. Pyun; Pyong K. Shin; Jin-Ho Seo

• Publisher: John Wiley and Sons
• Year: 1990
• Tongue: English
• Weight: 361 KB
• Volume: 36
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/bit.260360911

✦ Synopsis

Regulated promoters have advantages over nonregulated promoters in that gene expression levels can be controlled through manipulation of environmental conditions. The

Methods

Yeast Strain and Plasmid use of regulated promoters in the design of plasmid molecules allows separation of a cell growth stage from a cloned-gene expression period and hence minimizes competitive interactions between plasmids and host cells. This control property has been successfully implemented in various operating schemes to improve the productivity of recombinant cell fermentation processes. I4

The SUC2 gene system in Saccharomyces cerevisiae offers one means of environmental switching over plasmid protein synthesis. It provides a regulatory promoter, a transport signal sequence, and a structural gene encoding enzyme invertase. The expression of the yeast SUC2 promoter is repressed by a high glucose concentration in a growth medium, but derepressed by either a low glucose level or s u c r o ~e . ~

The yeast SUC2 gene system has been widely used in constructing recombinant plasmids for production of industrial proteins .6 Marten and Seo have studied the expression and transport of cloned invertase in a batch culture and compared quantitatively the secretion efficiency between the yeast SUC2 and the yeast a-mating factor signal sequences.'

In order to investigate the regulatory character of the yeast SUC2 promoter cloned on a multicopy plasmid in well-defined conditions, a continuous culture system was employed. Growth yield factors, specific ethanol productivity, and specific invertase activity as a function of dilution rates were determined for two different operating conditions: aerobic and anaerobic. In addition, specific invertase activity was correlated with residual glucose concentration to characterize the expression pattern of the cloned SUC2 gene in response to glucose levels in a fermentation medium.