Iterative algorithm-guided design of massive strain libraries, applied to itaconic acid production in yeast

Metabolic Engineering
Eric M YoungChristopher A Voigt

Abstract

Metabolic engineering requires multiple rounds of strain construction to evaluate alternative pathways and enzyme concentrations. Optimizing multigene pathways stepwise or by randomly selecting enzymes and expression levels is inefficient. Here, we apply methods from design of experiments (DOE) to guide the construction of strain libraries from which the maximum information can be extracted without sampling every possible combination. We use Saccharomyces cerevisiae as a host for a novel six-gene pathway to itaconic acid, selected by comparing alternative shunt pathways that bypass the mitochondrial TCA cycle. The pathway is distinctive for the use of acetylating acetaldehyde dehydrogenase to increase cytosolic acetyl-CoA pools, a bacterial enzyme to synthesize citrate in the cytosol, and an itaconic acid exporter. Precise control over the expression of each gene is enabled by a set of promoter-terminator pairs that span a 174-fold range. Two large combinatorial libraries (160 variants, 2.4 Mb and 32 variants, 0.6 Mb) are designed where the expression levels are selected by statistical methods (I-optimal response surface methodology, full factorial, or Plackett-Burman) with the intent of extracting different types of guiding in...Continue Reading

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Citations

Nov 30, 2018·Molecular Systems Biology·Felix MoserChristopher A Voigt
Nov 27, 2019·Proceedings of the National Academy of Sciences of the United States of America·Stefan BurénLuis M Rubio
Mar 13, 2020·Zeitschrift für Kinder- und Jugendpsychiatrie und Psychotherapie·Esther Strittmatter, Martin Holtmann
Mar 28, 2020·Applied Microbiology and Biotechnology·Xi ZhangYu Deng
Jan 25, 2019·Journal of Biological Engineering·Kealan ExleyRichard Ian Kitney
Dec 11, 2020·Metabolic Engineering·Rosanna YoungPaul S Freemont
Mar 7, 2021·Nature Communications·Joseph H CollinsEric M Young
Sep 25, 2018·Current Opinion in Biotechnology·Holly EkasHal S Alper
Apr 30, 2020·Metabolic Engineering Communications·Wan SunZengyi Shao
Jun 27, 2020·ACS Synthetic Biology·Mahsa BabaeiIrina Borodina
Apr 5, 2021·Metabolic Engineering·Miguel C Santoscoy, Laura R Jarboe
Aug 14, 2021·ACS Synthetic Biology·Jeanet ManteChris J Myers
Jan 9, 2019·Biochemistry·Zihe LiuJens Nielsen
Jan 8, 2021·ACS Synthetic Biology·James GilmanFilippo Menolascina
Sep 22, 2021·ACS Synthetic Biology·Jeanet ManteChris J Myers
Oct 8, 2021·Applied Microbiology and Biotechnology·Jinhua YangGuocheng Du

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