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The Saccharomyces cerevisiae pheromone-response is a metabolically active stationary phase for bio-production.

ABSTRACT: The growth characteristics and underlying metabolism of microbial production hosts are critical to the productivity of metabolically engineered pathways. Production in parallel with growth often leads to biomass/bio-product competition for carbon. The growth arrest phenotype associated with the Saccharomyces cerevisiae pheromone-response is potentially an attractive production phase because it offers the possibility of decoupling production from population growth. However, little is known about the metabolic phenotype associated with the pheromone-response, which has not been tested for suitability as a production phase. Analysis of extracellular metabolite fluxes, available transcriptomic data, and heterologous compound production (para-hydroxybenzoic acid) demonstrate that a highly active and distinct metabolism underlies the pheromone-response. These results indicate that the pheromone-response is a suitable production phase, and that it may be useful for informing synthetic biology design principles for engineering productive stationary phase phenotypes.

SUBMITTER: Williams TC

PROVIDER: S-EPMC5779721 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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The <i>Saccharomyces cerevisiae</i> pheromone-response is a metabolically active stationary phase for bio-production.

Williams Thomas C TC Peng Bingyin B Vickers Claudia E CE Nielsen Lars K LK

Metabolic engineering communications 20160511

The growth characteristics and underlying metabolism of microbial production hosts are critical to the productivity of metabolically engineered pathways. Production in parallel with growth often leads to biomass/bio-product competition for carbon. The growth arrest phenotype associated with the <i>Saccharomyces cerevisiae</i> pheromone-response is potentially an attractive production phase because it offers the possibility of decoupling production from population growth. However, little is known ...[more]

PMID: 29468120

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Project description:Saccharomyces cerevisiae is a versatile industrial host for chemical production and has been engineered to produce efficiently many valuable compounds. 2-Deoxy-scyllo-inosose (2-DOI) is an important precursor for the biosynthesis of 2-deoxystreptamine-containing aminoglycosides antibiotics and benzenoid metabolites. Bacterial and cyanobacterial strains have been metabolically engineered to generate 2-DOI; nevertheless, the production of 2-DOI using a yeast host has not been reported. Here, we have metabolically engineered a series of CEN.PK yeast strains to produce 2-DOI using a synthetically yeast codon-optimized btrC gene from Bacillus circulans. The expression of the 2-Deoxy-scyllo-inosose synthase (2-DOIS) gene was successfully achieved via an expression vector and through chromosomal integration at a high-expression locus. In addition, the production of 2-DOI was further investigated for the CEN.PK knockout strains of phosphoglucose isomerase (?pgi1), D-glucose-6-phosphate dehydrogenase (?zwf1) and a double mutant (?pgi1, ?zwf1) in a medium consisting of 2% fructose and 0.05% glucose as a carbon source. We have found that all the recombinant strains are capable of producing 2-DOI and reducing it into scyllo-quercitol and (-)-vibo-quercitol. Comparatively, the high production of 2-DOI and its analogs was observed for the recombinant CEN.PK-btrC carrying the multicopy btrC-expression vector. GC/MS analysis of culture filtrates of this strain showed 11 times higher response in EIC for the m/z 479 (methyloxime-tetra-TMS derivative of 2-DOI) than the YP-btrC recombinant that has only a single copy of btrC expression cassette integrated into the genomic DNA of the CEN.PK strain. The knockout strains namely ?pgi1-btrC and ?pgi1?zwf1-btrC, that are transformed with the btrC-expression plasmids, have inactive Pgi1 and produced only traces of the compounds. In contrast, ?zwf1-btrC recombinant which has intact pgi1 yielded relatively higher amount of the carbocyclic compounds. Additionally, 1H-NMR analysis of samples showed slow consumption of fructose and no accumulation of 2-DOI and the quercitols in the culture broth of the recombinant CEN.PK-btrC suggesting that S. cerevisiae is capable of assimilating 2-DOI.

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The Saccharomyces cerevisiae pheromone-response is a metabolically active stationary phase for bio-production.

Publications

The <i>Saccharomyces cerevisiae</i> pheromone-response is a metabolically active stationary phase for bio-production.

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