Organisms for the production of 1,3-butanediol

Patent metadata

Field	Value
Publication number	US-9708632-B2
Application number	US-201514673600-A
Country	US
Kind code	B2
Filing date	Mar 30, 2015
Priority date	Apr 30, 2009
Publication date	Jul 18, 2017
Grant date	Jul 18, 2017

Abstract

A non-naturally occurring microbial organism includes a microbial organism having a 1,3-butanediol (1,3-BDO) pathway having at least one exogenous nucleic acid encoding a 1,3-BDO pathway enzyme expressed in a sufficient amount to produce 1,3-BDO. The pathway includes an enzyme selected from a 2-amino-4-ketopentanoate (AKP) thiolase, an AKP dehydrogenase, a 2-amino-4-hydroxypentanoate aminotransferase, a 2-amino-4-hydroxypentanoate oxidoreductase (deaminating), a 2-oxo-4-hydroxypentanoate decarboxylase, a 3-hydroxybutyraldehyde reductase, an AKP aminotransferase, an AKP oxidoreductase (deaminating), a 2,4-dioxopentanoate decarboxylase, a 3-oxobutyraldehyde reductase (ketone reducing), a 3-oxobutyraldehyde reductase (aldehyde reducing), a 4-hydroxy-2-butanone reductase, an AKP decarboxylase, a 4-aminobutan-2-one aminotransferase, a 4-aminobutan-2-one oxidoreductase (deaminating), a 4-aminobutan-2-one ammonia-lyase, a butenone hydratase, an AKP ammonia-lyase, an acetylacrylate decarboxylase, an acetoacetyl-CoA reductase (CoA-dependent, aldehyde forming), an acetoacetyl-CoA reductase (CoA-dependent, alcohol forming), an acetoacetyl-CoA reductase (ketone reducing), a 3-hydroxybutyryl-CoA reductase (aldehyde forming), a 3-hydroxybutyryl-CoA reductase (alcohol forming), a 4-hydroxybutyryl-CoA dehydratase, and a crotonase. A method for producing 1,3-BDO, includes culturing such microbial organisms under conditions and for a sufficient period of time to produce 1,3-BDO.

First claim

What is claimed is:

1. A method for producing 1,3-BDO comprising culturing a non-naturally occurring microbial organism under conditions and for a sufficient period of time to produce 1,3-BDO; said non-naturally occurring microbial organism comprising at least two exogenous nucleic acids each encoding a 1,3-BDO pathway enzyme expressed in a sufficient amount to produce 1,3-BDO; said 1,3-BDO pathway enzyme comprising
an acetoacetyl-CoA reductase (ketone reducing) that converts acetoacetyl-CoA to 3-hydroxybutyryl-CoA; and
(i) a 3-hydroxybutyryl-CoA reductase (aldehyde forming) that converts 3-hydroxybutyryl-CoA to 3-hydroxybutyraldehyde; or
(ii) a 3-hydroxybutyraldehyde reductase that converts 3-hydroxybutyraldehyde to 1,3-butanediol.

2. The method of claim 1 , wherein the at least two exogenous nucleic acids encode the acetoacetyl-CoA reductase (ketone reducing) and the 3-hydroxybutyryl-CoA reductase (aldehyde forming).

3. The method of claim 1 , wherein the at least two exogenous nucleic acids encode the acetoacetyl-CoA reductase (ketone reducing) and the 3-hydroxybutyraldehyde reductase.

4. The method of claim 1 , wherein the non-naturally occurring microbial organism comprising three exogenous nucleic acids each encoding (1) an acetoacetyl-CoA reductase (ketone reducing); (2) a 3-hydroxybutyryl-CoA reductase (aldehyde forming); and (3) a 3-hydroxybutyraldehyde reductase.

5. The method of claim 1 , wherein said acetoacetyl-CoA reductase (ketone reducing) is encoded by one or more genes selected from the group consisting of thrA, akthr2, hom6, hom1, hom2, fadB, fadJ, Hbd2, Hbd1, hbd, HSD 17B10, phbB, phaB, Msed_1423, Msed_0399, Msed_0389,Msed_1993, adh, adhA, adh-A, mdh, ldhA, ldh, and bdh.

6. The method of claim 1 , wherein said 3-hydroxybutyryl-CoA reductase (aldehyde forming) is encoded by one or more genes selected from the group consisting of acr1, sucD, bphG, bld, adhE, Msed_0709, mcr, asd-2, Saci_2370, Ald, and eutE.

7. The method of claim 1 , wherein said 3-hydroxybutyraldehdye reductase is encoded by one or more genes selected from the group consisting of alrA, ADH2, yqhD, bdh I, bdh II, adhA, 4hbd, adhI, P84067, mmsb, dhat, and 3hidh.

8. The method of claim 1 , wherein said microbial organism is in a substantially anaerobic culture medium.

9. The method of claim 1 , wherein at least one exogenous nucleic acid is a heterologous nucleic acid.

10. The method of claim 1 further comprising separating 1,3-BDO from other components in the culture.

11. The method of claim 10 , wherein the separating comprises extraction, continuous liquid-liquid extraction, pervaporation, membrane filtration, membrane separation, reverse osmosis, electrodialysis, distillation, crystallization, centrifugation, extractive filtration, ion exchange chromatography, absorption chromatography, or ultrafiltration.

12. The method of claim 10 , wherein the separating comprising distillation.

Assignees

• Genomatica Inc

Inventors

• Burgard Anthony P
• Burk Mark J
• Osterhout Robin E
• Pharkya Priti

Classifications

• C12Y403/01

  Ammonia-lyases (4.3.1) · CPC title

• C12N1/02

  Separating microorganisms from their culture media · CPC title

• C12Y206/01

  Transaminases (2.6.1) · CPC title

• C12N9/1096

  transferring nitrogenous groups (2.6) · CPC title

• C12N9/0006 Primary

  acting on CH-OH groups as donors (1.1) · CPC title