Primary alcohol producing organisms
View Patent ↗The invention provides a non-naturally occurring microbial organism having at least one exogenous gene insertion and/or one or more gene disruptions that confer production of primary alcohols. A method for producing long chain alcohols includes culturing these non-naturally occurring microbial organisms.
1. A non-naturally occurring microbial organism, comprising exogenous nucleic acids encoding a malonyl-CoA-independent fatty acid synthesis (FAS) pathway and exogenous nucleic acids encoding an acyl-reduction pathway expressed in sufficient amounts to produce a primary alcohol having 4-24 carbon atoms, said malonyl-CoA-independent FAS pathway comprising exogenous nucleic acids encoding ketoacyl-CoA acyltransferase or ketoacyl-CoA thiolase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase and enoyl-CoA reductase, said acyl-reduction pathway comprising one or more exogenous nucleic acids encoding an acyl-CoA reductase and an alcohol dehydrogenase, wherein said exogenous nucleic acids encoding said malonyl-CoA-independent FAS pathway and said exogenous nucleic acids encoding said acyl-reduction pathway are expressed in the cytosol, wherein said ketoacyl-CoA acyltransferase or ketoacyl-CoA thiolase converts acyl-CoA to β-ketoacyl-CoA, wherein said 3-hydroxyacyl-CoA dehydrogenase converts β-ketoacyl-CoA to β-hydroxyacyl-CoA, wherein said enoyl-CoA hydratase converts β-hydroxyacyl-CoA to trans-2-enoyl-CoA, wherein said enoyl-CoA reductase converts trans-2-enoyl-CoA to acyl-CoA, wherein said acyl-CoA reductase converts acyl-CoA to an aldehyde, wherein said alcohol dehydrogenase converts an aldehyde to said primary alcohol.
2. The non-naturally occurring microbial organism of claim 1 , wherein said exogenous nucleic acids encoding said malonyl-CoA-independent FAS pathway comprise ketoacyl-CoA acyltransferase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase and enoyl-CoA reductase.
3. The non-naturally occurring microbial organism of claim 1 , wherein said exogenous nucleic acids encoding said malonyl-CoA-independent FAS pathway comprise ketoacyl-CoA thiolase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase and enoyl-CoA reductase.
4. The non-naturally occurring microbial organism of claim 1 , wherein said one or more exogenous nucleic acids encoding an acyl-reduction pathway encode an enzyme comprising acyl-CoA reductase and alcohol dehydrogenase activity, wherein said enzyme converts acyl-CoA to said primary alcohol.
5. The non-naturally occurring microbial organism of claim 4 , wherein said enzyme having acyl-CoA reductase and alcohol dehydrogenase activity comprises fatty alcohol forming acyl-CoA reductase (FAR).
6. The non-naturally occurring microbial organism of claim 1 , wherein at least one of said exogenous nucleic acid further comprises a heterologous encoding nucleic acid.
7. The non-naturally occurring microbial organism of claim 1 , wherein said primary alcohol is produced in amounts at least 10% greater levels compared to a microbial organism lacking said exogenous nucleic acid encoding a malonyl-CoA-independent FAS pathway and exogenous nucleic acids encoding an acyl-reduction pathway.
8. The non-naturally occurring microbial organism of claim 1 , further comprising a substantially anaerobic culture medium.
9. The non-naturally occurring microbial organism of claim 1 , wherein said alcohol having between 4-24 carbon atoms is selected from butanol, hexanol, heptanol, octanol, nonanol, decanol, dodecanol, tetradecanol or hexadecanol.
10. A method for producing a primary alcohol, comprising culturing the non-naturally occurring microbial organism of claim 1 , wherein the malonyl-CoA-independent fatty acid synthesis (FAS) pathway and the acyl-reduction pathway enzymes are expressed in sufficient amounts to produce a primary alcohol having 4-24 carbon atoms under substantially anaerobic conditions for a sufficient period of time to produce said primary alcohol.
11. The method of claim 10 , wherein said exogenous nucleic acid encoding an acyl-reduction pathway enzyme comprises an enzyme having acyl-CoA reductase and alcohol dehydrogenase activity, wherein said enzyme converts acyl-CoA to said primary alcohol.
12. The method of claim 11 , wherein said enzyme having acyl-CoA reductase and alcohol dehydrogenase activity comprises fatty alcohol forming acyl-CoA reductase (FAR).
13. The method of claim 10 , wherein at least one exogenous nucleic acid is a heterologous encoding nucleic acid.
14. The method of claim 10 , wherein said primary alcohol is produced in amounts at least 10% greater levels compared to a microbial organism lacking said exogenous nucleic acids encoding said malonyl-CoA-independent FAS pathway enzymes.
15. The method of claim 10 , wherein said primary alcohol is selected from butanol, hexanol, heptanol, octanol, nonanol, decanol, dodecanol, tetradecanol or hexadecanol.
16. The method of claim 10 , further comprising isolating said primary alcohol.