Genetic engineering to enhance the Ehrlich pathway and alter carbon flux for increased isobutanol production from glucose by Saccharomyces cerevisiae

► We engineered Saccharomyces cerevisiae for increased isobutanol production. ► Ehrlich pathway was enhanced by overexpressing suitable KDC and ADH enzymes. ► Ethanol flux was altered by combining ILV2 overexpression and PDC1 disruption. The production of higher alcohols by engineered bacteria has r...

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Bibliographic Details
Published in:Journal of biotechnology 2012-05, Vol.159 (1-2), p.32-37
Main Authors: Kondo, Takashi, Tezuka, Hironori, Ishii, Jun, Matsuda, Fumio, Ogino, Chiaki, Kondo, Akihiko
Format: Article
Language:English
Subjects:
acid tolerance
Aerobiosis
alcohol dehydrogenase
Alcohol Dehydrogenase - genetics
Alcohol Dehydrogenase - metabolism
biosynthesis
biotechnology
Butanols - analysis
Butanols - metabolism
carbon
Carbon - metabolism
Carbon flux
Carboxy-Lyases - genetics
Carboxy-Lyases - metabolism
Ehrlich pathway
engineering
ethanol
Ethanol - analysis
Ethanol - metabolism
Fermentation
gene overexpression
genes
genetic engineering
Genetic Engineering - methods
genetically engineered microorganisms
glucose
Glucose - metabolism
Hydro-Lyases - genetics
Hydro-Lyases - metabolism
Isobutanol
Metabolic Networks and Pathways - genetics
pyruvate decarboxylase
Pyruvate Decarboxylase - genetics
Pyruvate Decarboxylase - metabolism
pyruvic acid
Saccharomyces cerevisiae
Saccharomyces cerevisiae - enzymology
Saccharomyces cerevisiae - genetics
Saccharomyces cerevisiae - metabolism
Saccharomyces cerevisiae Proteins - genetics
Saccharomyces cerevisiae Proteins - metabolism
valine
yeasts
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Summary:► We engineered Saccharomyces cerevisiae for increased isobutanol production. ► Ehrlich pathway was enhanced by overexpressing suitable KDC and ADH enzymes. ► Ethanol flux was altered by combining ILV2 overexpression and PDC1 disruption. The production of higher alcohols by engineered bacteria has received significant attention. The budding yeast, Saccharomyces cerevisiae, has considerable potential as a producer of higher alcohols because of its capacity to naturally fabricate fusel alcohols, in addition to its robustness and tolerance to low pH. However, because its natural productivity is not significant, we considered a strategy of genetic engineering to increase production of the branched-chain higher alcohol isobutanol, which is involved in valine biosynthesis. Initially, we overexpressed 2-keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH) in S. cerevisiae to enhance the endogenous activity of the Ehrlich pathway. We then overexpressed Ilv2, which catalyzes the first step in the valine synthetic pathway, and deleted the PDC1 gene encoding a major pyruvate decarboxylase with the intent of altering the abundant ethanol flux via pyruvate. Through these engineering steps, along with modification of culture conditions, the isobutanol titer of S. cerevisiae was elevated 13-fold, from 11mg/l to 143mg/l, and the yield was 6.6mg/g glucose, which is higher than any previously reported value for S. cerevisiae.
ISSN:0168-1656
1873-4863
DOI:10.1016/j.jbiotec.2012.01.022