Print Email Facebook Twitter Elimination of aromatic fusel alcohols as by-products of Saccharomyces cerevisiae strains engineered for phenylpropanoid production by 2-oxo-acid decarboxylase replacement Title Elimination of aromatic fusel alcohols as by-products of Saccharomyces cerevisiae strains engineered for phenylpropanoid production by 2-oxo-acid decarboxylase replacement Author Else-Hassing, J. (TU Delft BT/Industriele Microbiologie) Buijs, Joran (Student TU Delft) Blankerts, Nikki (Student TU Delft) Luttik, M.A.H. (TU Delft BT/Industriele Microbiologie) de Hulster, A.F. (TU Delft BT/Industriele Microbiologie) Pronk, J.T. (TU Delft BT/Biotechnologie) Daran, J.G. (TU Delft BT/Industriele Microbiologie) Department BT/Biotechnologie Date 2021 Abstract Engineered strains of the yeast Saccharomyces cerevisiae are intensively studied as production platforms for aromatic compounds such as hydroxycinnamic acids, stilbenoids and flavonoids. Heterologous pathways for production of these compounds use L-phenylalanine and/or L-tyrosine, generated by the yeast shikimate pathway, as aromatic precursors. The Ehrlich pathway converts these precursors to aromatic fusel alcohols and acids, which are undesirable by-products of yeast strains engineered for production of high-value aromatic compounds. Activity of the Ehrlich pathway requires any of four S. cerevisiae 2-oxo-acid decarboxylases (2-OADCs): Aro10 or the pyruvate-decarboxylase isoenzymes Pdc1, Pdc5, and Pdc6. Elimination of pyruvate-decarboxylase activity from S. cerevisiae is not straightforward as it plays a key role in cytosolic acetyl-CoA biosynthesis during growth on glucose. In a search for pyruvate decarboxylases that do not decarboxylate aromatic 2-oxo acids, eleven yeast and bacterial 2-OADC-encoding genes were investigated. Homologs from Kluyveromyces lactis (KlPDC1), Kluyveromyces marxianus (KmPDC1), Yarrowia lipolytica (YlPDC1), Zymomonas mobilis (Zmpdc1) and Gluconacetobacter diazotrophicus (Gdpdc1.2 and Gdpdc1.3) complemented a Pdc− strain of S. cerevisiae for growth on glucose. Enzyme-activity assays in cell extracts showed that these genes encoded active pyruvate decarboxylases with different substrate specificities. In these in vitro assays, ZmPdc1, GdPdc1.2 or GdPdc1.3 had no substrate specificity towards phenylpyruvate. Replacing Aro10 and Pdc1,5,6 by these bacterial decarboxylases completely eliminated aromatic fusel-alcohol production in glucose-grown batch cultures of an engineered coumaric acid-producing S. cerevisiae strain. These results outline a strategy to prevent formation of an important class of by-products in ‘chassis’ yeast strains for production of non-native aromatic compounds. Subject Ehrlich pathwayFusel alcoholsPhenylpropanoidPhenylpyruvate decarboxylasePyruvate decarboxylaseSaccharomyces cerevisiae To reference this document use: http://resolver.tudelft.nl/uuid:2f86b655-6641-471b-90e2-9000485b5570 DOI https://doi.org/10.1016/j.mec.2021.e00183 ISSN 2214-0301 Source Metabolic Engineering Communications, 13 Part of collection Institutional Repository Document type journal article Rights © 2021 J. Else-Hassing, Joran Buijs, Nikki Blankerts, M.A.H. Luttik, A.F. de Hulster, J.T. Pronk, J.G. Daran Files PDF 1_s2.0_S2214030121000237_main.pdf 4.38 MB Close viewer /islandora/object/uuid:2f86b655-6641-471b-90e2-9000485b5570/datastream/OBJ/view