Investigating strain dependency in the production of aromatic compounds in Saccharomyces cerevisiae. Issue 12 (30th June 2016)

• Record Type: Journal Article
• Authors: Suástegui, Miguel; Guo, Weihua; Feng, Xueyang; Shao, Zengyi
• Is Part Of: Biotechnology and bioengineering. Volume 113:Issue 12(2016)
• Page Start: 2676
• Page End: 2685
• Publication Date: 2016-06-30
• Subjects: shikimate pathway -- protein overexpression burden -- pathway balancing -- 13C‐metabolic flux analysis; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.26037
• Languages: English
• ISSNs: 0006-3592

Abstract

ABSTRACT: Although Saccharomyces cerevisiae is the most highly domesticated yeast, strain dependency in biotechnological processes still remains as a common, yet poorly understood phenomenon. To investigate this, the entrance to the aromatic amino acid biosynthetic pathway was compared in four commonly used S. cerevisiae laboratory strains. The strains were engineered to accumulate shikimate by overexpressing a mutant version of the pentafunctional ARO1 enzyme with disrupted activity in the shikimate kinase subunit. Carbon tracing and 13 C metabolic flux analysis combined with quantitative PCR, revealed that precursor availability and shikimate production were dramatically different in the four equally engineered strains, which were found to be correlated with the strains' capacity to deal with protein overexpression burden. By implementing a strain‐dependent approach, the genetic platform was reformulated, leading to an increase in yield and titer in all strains. The highest producing strain, INVSc1‐SA3, produced 358 mg L −1 of shikimate with a yield of 17.9 mg g −1 glucose. These results underline the importance of strain selection in developing biological manufacturing processes, demonstrate the first case of high production of shikimate in yeast, and provide an appropriate platform for strain selection for future production of aromatic compounds. Biotechnol. Bioeng. 2016;113: 2676–2685. © 2016 Wiley Periodicals, Inc. Abstract : A metabolic engineering strategy was designed based on differential ability of four Saccharomyces cerevisiae strains to produce the aromatic amino acid intermediate shikimate. By understanding the factors that caused low production in the strains, the plasmid platform was simplified and reconstructed, therefore enabling higher accumulation of shikimate in the four engineered strains.