Biofuel production by recombinant microorganisms
AssigneeThe Regents of the University of California
InventorsJames C. Liao, Shota Atsumi, Kevin M. Smith, Roa Pu Claire Shen, Anthony F. Cann, Michael R. Connor
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Abstract
Provided herein are metabolically-modified microorganisms useful for producing biofuels. More specifically, provided herein are methods of producing high alcohols including isobutanol, 1-butanol, 1-propanol, 2-methyl-1-butanol, 3-methyl-1-butanol and 2-phenylethanol from a suitable substrate.
Claims12
A recombinant E. coli comprising:
- (a) an acetolactate synthase polypeptide having at least 90% identity to SEQ ID NO:82 and having acetolactate synthase activity;
- (b) an acetohydroxy acid isomeroreductase polypeptide having at least 90% identity to SEQ ID NO:44 and having acetohydroxy acid isomeroreductase activity;
- (c) a dihydroxy-acid dehydratase polypeptide having at least 90% identity to SEQ ID NO:46 and having dihvdroxv-acid dehydratase activity;
- (d) a 2-keto-acid decarboxylase polypeptide having at least 90% identity to SEQ ID NO:28 and having 2-keto acid decarboxylase activity, wherein the 2-keto-acid decarboxylase can convert α-keto-isovalerate to Isobutyraldehyde;
- and
- (e) a NADH-dependent alcohol dehydrogenase having at least 90% identity to SEQ ID NO:38 and that converts isobutyraldehyde to isobutanol,
- and having a disruption of an endogenous adhE gene, or the endogenous adhE gene and an endogenous IdhA gene;
- and
- wherein the recombinant E. coli produces isobutanol.
A recombinant E. coli comprising a genotype for overexpressing:
- (a) a naturally occurring bacterial gene encoding an acetolactate synthase or variants of a Bacillus subtilis AlsS having at least 90% identity to SEQ ID NO:82 and having acetolactate synthase activity;
- (b) a naturally occurring bacterial gene encoding an acetohydroxy acid isomeroreductase or variants of a E. coli ilvC having at least 90% identity to SEQ ID NO:44 and having acetohydroxy acid isomeroreductase activity;
- (c) a naturally occurring bacterial gene encoding a dihydroxy-acid dehydratase or variants of an E. coli ilvD having at least 90% identity to SEQ ID NO:46 and having dihydroxy-acid dehydratase activity;
- and
- (d) a Lactococcus lactis gene encoding a 2-keto-acid decarboxylase or variants of a Lactococcus lactis KivD having at least 90% identity to SEQ ID NO:28 and converts α-keto-isovalerate to Isobutvraldehyde;
- (e) a Saccharomyces cerevisiae ADH2 gene encoding a NADH-dependent alcohol dehydrogenase that converts isobutyraldehyde to isobutanol or variants of a Saccharomyces cerevisiae adh2 having at least 90% identity to SEQ ID NO:38 and that converts Isobutyraldehyde to isobutanol;
- wherein the recombinant E. coli comprises reduced expression or activity of an endogenous adhE gene or the endogenous adhE gene and an endogenous IdhA gene, and
- wherein the recombinant E. coli produces isobutanol.
A recombinant E. coli that produces isobutanol, wherein isobutanol is produced from a metabolite comprising a 2-keto acid and wherein said 2-keto acid is converted to isobutanol by a 2-keto acid decarboxylase and a NADH-dependent alcohol dehydrogenase;
- wherein said 2-keto acid decarboxylase is encoded by a Lactococcus lactis KivD gene or variants of a Lactococcus lactis KivD having at least 90% identity to SEQ ID NO:28 that converts α-keto-isovalerate to Isobutyraldehyde;
- and wherein the NADH-dependent alcohol dehydrogenase is encoded by a Saccharomyces cerevisiae adh2 gene or variants of a Saccharomyces cerevisiae adh2 having at least 90% identity to SEQ ID NO:38 and converts Isobutyraldehyde to isobutanol;
- and wherein the recombinant E. coli has been engineered to disrupt an ethanol dehydrogenase encoded by an endogenous adhE gene or the endogenous adhE gene and an endogenous IdhA gene.
A recombinant microorganism designated SA237 and having ATTC Patent Deposit Designation PTA-8945.
Dependencies are inferred from the claim text.