An engineered fatty acid synthase combined with a carboxylic acid reductase enables de novo production of 1-octanol in Saccharomyces cerevisiae

  • Background: The ideal biofuel should not only be a regenerative fuel from renewable feedstocks, but should also be compatible with the existing fuel distribution infrastructure and with normal car engines. As the so-called drop-in biofuel, the fatty alcohol 1-octanol has been described as a valuable substitute for diesel and jet fuels and has already been produced fermentatively from sugars in small amounts with engineered bacteria via reduction of thioesterase-mediated premature release of octanoic acid from fatty acid synthase or via a reversal of the β-oxidation pathway. Results: The previously engineered short-chain acyl-CoA producing yeast Fas1R1834K/Fas2 fatty acid synthase variant was expressed together with carboxylic acid reductase from Mycobacterium marinum and phosphopantetheinyl transferase Sfp from Bacillus subtilis in a Saccharomyces cerevisiae Δfas1 Δfas2 Δfaa2 mutant strain. With the involvement of endogenous thioesterases, alcohol dehydrogenases, and aldehyde reductases, the synthesized octanoyl-CoA was converted to 1-octanol up to a titer of 26.0 mg L−1 in a 72-h fermentation. The additional accumulation of 90 mg L−1 octanoic acid in the medium indicated a bottleneck in 1-octanol production. When octanoic acid was supplied externally to the yeast cells, it could be efficiently converted to 1-octanol indicating that re-uptake of octanoic acid across the plasma membrane is not limiting. Additional overexpression of aldehyde reductase Ahr from Escherichia coli nearly completely prevented accumulation of octanoic acid and increased 1-octanol titers up to 49.5 mg L−1. However, in growth tests concentrations even lower than 50.0 mg L−1 turned out to be inhibitory to yeast growth. In situ extraction in a two-phase fermentation with dodecane as second phase did not improve growth, indicating that 1-octanol acts inhibitive before secretion. Furthermore, 1-octanol production was even reduced, which results from extraction of the intermediate octanoic acid to the organic phase, preventing its re-uptake. Conclusions: By providing chain length control via an engineered octanoyl-CoA producing fatty acid synthase, we were able to specifically produce 1-octanol with S. cerevisiae. Before metabolic engineering can be used to further increase product titers and yields, strategies must be developed that cope with the toxic effects of 1-octanol on the yeast cells.

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Author:Sandra Henritzi, Manuel Fischer, Martin GriningerORCiDGND, Igor-Mislav OrebORCiDGND, Eckhard Boles
URN:urn:nbn:de:hebis:30:3-466075
DOI:https://doi.org/10.1186/s13068-018-1149-1
ISSN:1754-6834
Pubmed Id:https://pubmed.ncbi.nlm.nih.gov/29881455
Parent Title (English):Biotechnology for biofuels
Publisher:BioMed Central
Place of publication:London
Document Type:Article
Language:English
Year of Completion:2018
Date of first Publication:2018/06/01
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2018/06/14
Tag:1-octanol; Biofuel; Caprylic acid; Carboxylic acid reductase; Fatty acid synthase; Fatty alcohol; Octanoic acid; Saccharomyces cerevisiae; Short-chain fatty acids; Yeast
Volume:11
Issue:Art. 150
Page Number:12
First Page:1
Last Page:12
Note:
Open Access: This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
HeBIS-PPN:433867051
Institutes:Biochemie, Chemie und Pharmazie / Biochemie und Chemie
Biowissenschaften / Biowissenschaften
Exzellenzcluster / Exzellenzcluster Makromolekulare Komplexe
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Sammlungen:Universitätspublikationen
Open-Access-Publikationsfonds:Biowissenschaften
Licence (German):License LogoCreative Commons - Namensnennung 4.0