Method for Acetate Consumption During Ethanolic Fermentaion of Cellulosic Feedstocks
The present invention provides for novel metabolic pathways to detoxify biomass-derived acetate via metabolic conversion to ethanol, acetone, or isopropanol. More specifically, the invention provides for a recombinant microorganism comprising one or more native and/or heterologous enzymes that function in one or more first engineered metabolic pathways to achieve: (1) conversion of acetate to ethanol; (2) conversion of acetate to acetone; or (3) conversion of acetate to isopropanol; and one or more native and/or heterologous enzymes that function in one or more second engineered metabolic pathways to produce an electron donor used in the conversion of acetate to less inhibitory compounds; wherein the one or more native and/or heterologous enzymes is activated, upregulated, or downregulated.
1 . A recombinant yeast microorganism comprising an engineered metabolic pathway to convert acetate to ethanol comprising
(i) the conversion of acetate to acetyl-CoA and
(ii) the conversion of acetyl-CoA to ethanol,
wherein said microorganism expresses a heterologous enzyme that is an NADPH-specific alcohol dehydrogenase and wherein said microorganism expresses a native and/or heterologous enzyme that is encoded by a zwf1 polynucleotide recombinantly introduced into said microorganism.
2 . The recombinant microorganism of claim 1 , wherein said NADPH-specific alcohol dehydrogenase is from a microorganism selected from the group consisting of T. pseudethanolicus, C. beijerinckii, Entamoeba histolytica, Cucumis melo , and S. cerevisiae.
3 . The recombinant microorganism of claim 1 , wherein said acetate is converted to acetyl-CoA by an acetyl-CoA transferase (ACS).
4 . The recombinant microorganism of claim 1 , wherein said acetate is converted to acetyl-P by an acetate kinase; and wherein said acetyl-P is converted to acetyl-CoA by a phosphotransacetylase.
5 . The recombinant microorganism of claim 1 , wherein said acetyl-CoA is converted to acetaldehyde by an acetaldehyde dehydrogenase; and wherein said acetaldehyde is converted to ethanol by an alcohol dehydrogenase.
6 . The recombinant microorganism of claim 1 , wherein said acetyl-CoA is converted to ethanol by a bifunctional acetaldehyde/alcohol dehydrogenase.
7 . A process for converting biomass to ethanol comprising contacting biomass with a recombinant microorganism according to claim 1 .
8 . An engineered metabolic pathway for reducing or removing acetate from consolidated bioprocessing (CBP) media according to claim 1 .
9 . A recombinant yeast microorganism comprising an engineered metabolic pathway to convert acetate to ethanol comprising
(i) the conversion of acetate to acetyl-CoA and
(ii) the conversion of acetyl-CoA to ethanol,
wherein said microorganism expresses a heterologous enzyme that is an NADPH-specific alcohol dehydrogenase and wherein said microorganism expresses a native and/or heterologous enzyme that is an acetyl-CoA synthetase recombinantly introduced into said microorganism.
10 . The recombinant microorganism of claim 9 , wherein said NADPH-specific alcohol dehydrogenase is from a microorganism selected from the group consisting of T. pseudethanolicus, C. beijerinckii, Entamoeba histolytica, Cucumis melo , and S. cerevisiae.
11 . The recombinant microorganism of claim 9 , wherein said acetyl-CoA synthetase is from Saccharomyces cerevisiae, Saccharomyces kluyveri, Zygosaccharomyces bailii , or Acetobacter aceti.
12 . A recombinant yeast microorganism comprising an engineered metabolic pathway to convert acetate to ethanol comprising
(i) the conversion of acetate to acetyl-CoA and
(ii) the conversion of acetyl-CoA to ethanol,
wherein said microorganism expresses a heterologous enzyme that is an NADPH-specific alcohol dehydrogenase and wherein said microorganism expresses a native and/or heterologous enzyme that is an NADH-specific alcohol dehydrogenase recombinantly introduced into said microorganism.
13 . The recombinant microorganism of claim 12 , wherein said NADPH-specific alcohol dehydrogenase is from a microorganism selected from the group consisting of T. pseudethanolicus, C. beijerinckii, Entamoeba histolytica, Cucumis melo , and S. cerevisiae.
14 . A recombinant yeast microorganism comprising an engineered metabolic pathway to convert acetate to ethanol comprising
(i) the conversion of acetate to acetyl-CoA and
(ii) the conversion of acetyl-CoA to ethanol,
wherein said microorganism expresses a heterologous enzyme that is an NADPH-specific alcohol dehydrogenase and wherein a native NADH-specific alcohol dehydrogenase in said microorganism is down-regulated.
15 . The recombinant microorganism of claim 14 , wherein said NADPH-specific alcohol dehydrogenase is from a microorganism selected from the group consisting of T. pseudethanolicus, C. beijerinckii, Entamoeba histolytica, Cucumis melo , and S. cerevisiae.
16 . A recombinant yeast microorganism comprising an engineered metabolic pathway to convert acetate to ethanol comprising
(i) the conversion of acetate to acetyl-CoA and
(ii) the conversion of acetyl-CoA to ethanol,
wherein said microorganism
(a) expresses a native and/or heterologous enzyme that is an NADH-dependent acetaldehyde dehydrogenase recombinantly introduced into said microorganism,
(b) expresses a native and/or heterologous enzyme that is an NADPH-specific xylose reductase recombinantly introduced into said microorganism, and
(c) expresses a native and/or heterologous enzyme that is an NADH-specific xylitol dehydrogenase recombinantly introduced into said microorganism.