The effects of residual catabolite repression and the importance of induction timing were determined for a temperature-sensitive (ts) GAL-regulated stable yeast expression system. The Saccharomyces cerevisiae strain employed carries a reg1 mutation inhibiting catabolite repression, and a ts mutation enabling induction of the regulated GAL promoters by a temperature shift to 35 degrees C. Despite the reg1 mutation and induction method, glucose depressed lacZ expression from a GAL1 promoter during batch culture. beta-Galactosidase specific activity was consistently lower at higher initial glucose concentrations in both SDC (semi-defined) and YPDa (complex) media; decreases of 18-36% were observed as glucose concentration was increased between 1, 3, 5, and 10 g l-1. However, the reductions in beta-galactosidase specific activity due to residual catabolite repression were more than balanced by substantial improvements in biomass yield at higher glucose levels. Therefore, productivity rose with increasing glucose concentration; in YPDa medium, increasing initial glucose from 1 to 10 g l-1 resulted in a 2.6-fold increase in beta-galactosidase volumetric activity. Due to the negative effects of shifting temperature to 35 degrees C, the trade-offs between optimum growth and a lengthy induction period were also evaluated. Delaying the time of induction reduced final specific activities but improved cell yield, and waiting 14 h into batch culture to induce lacZ expression provided modest 9-15% improvements in overall productivity.

• PMID: 7764717
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Journal Article | Research Support, U.S. Gov't, Non-P.H.S.

Authors

Napp SJ, Da Silva NA

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