Use of enzymes which catalyze pyruvate synthesis from formate and acetyl-CoA and bacteria expressing same

Patent metadata

Field	Value
Publication number	US-10053712-B2
Application number	US-201415101097-A
Country	US
Kind code	B2
Filing date	Dec 10, 2014
Priority date	Dec 10, 2013
Publication date	Aug 21, 2018
Grant date	Aug 21, 2018

Abstract

An isolated microorganism is disclosed being genetically modified to express pyruvate formate lyase (PFL) or 2-ketobutyrate formate lyase, wherein acetyl-CoA of the microorganism is converted to pyruvate in the presence of formate in a single step reaction, wherein the net flux of the reaction is in the direction of pyruvate synthesis.
Uses of the microorganism and products comprising same are also disclosed.

First claim

What is claimed is:

1. An isolated microorganism that is a bacterium or yeast, that is genetically modified to express a pyruvate formate lyase (PFL) or to express a 2-ketobutyrate formate lyase, wherein the microorganism further expresses a polypeptide which activates said PFL, wherein the microorganism does not express isocytrate lyase (aceA) and or malate synthase, wherein the microorganism is capable of converting acetyl CoA to pyruvate, wherein the source of said acetyl coA is:
(i) internal metabolism; or
(ii) externally supplied acetate in a medium,
wherein said medium comprises an amount of formate and/or acetate which favors the net flux of the reaction to be in the direction of pyruvate synthesis.

2. The isolated microorganism of claim 1 , wherein said pyruvate formate lyase is encoded by a gene selected from the group consisting of pflB, pflD and ybiW.

3. The microorganism of claim 1 , that is capable of generating an internal source of acetyl-CoA.

4. The microorganism of claim 1 , that is formatotrophic.

5. The microorganism of claim 3 , that expresses at least one enzyme selected from the group consisting of pyruvate carboxylase, malate dehydrogenase, malate-CoA ligase and malyl-CoA lyase.

6. The microorganism of claim 1 , that is a bacterium.

7. The microorganism of claim 6 , wherein said bacterium comprises Escherichia.

8. The microorganism of claim 1 , that is incapable of transporting and utilizing sugar.

9. The microorganism of claim 8 , not expressing at least one of the enzymes selected from the group consisting of phosphoglyceromutase, isocitrate lyase and phosphogluconate dehydratase.

10. The microorganism of claim 1 , that is a yeast.

11. The microorganism of claim 1 , capable of producing a biofuel.

12. A system for culturing the microorganism of claim 4 and an electrode for providing electrons to generate formate from CO 2 .

13. A cell culture comprising the isolated microorganism of claim 1 and a medium suitable for propagation of the microorganism.

14. A cell culture, comprising an isolated microorganism being genetically modified to express an enzyme of a reaction which converts acetyl-CoA of the microorganism to pyruvate in the presence of formate in a single step, and a medium comprising an amount of acetate and/or formate which favors the net flux of the reaction to be in the direction of pyruvate synthesis.

15. The cell culture of claim 13 , wherein the medium comprises an amount of acetate and/or formate which favors the net flux of the reaction to be in the direction of pyruvate synthesis.

16. A method of generating a biofuel comprising culturing the microorganism of claim 11 under conditions that allow for biofuel formation, thereby generating the biofuel.

17. A method of scavenging acetate from an acetate-containing medium comprising contacting the microorganism of claim 1 , with said medium, thereby scavenging the acetate from the acetate-containing medium.

18. A method of scavenging formate from a formate-containing medium comprising contacting the microorganism of claim 1 with said medium, thereby scavenging the formate from the formate-containing medium.

Assignees

• Yeda Res & Dev

Inventors

• Bar-Even Arren
• Milo Ron
• Noor Elad
• Zelcbuch Lior

Classifications

• C12P7/16

  Butanols · CPC title

• C12P5/007

  containing one or more isoprene units, i.e. terpenes (carotenes C12P23/00) · CPC title

• C12P5/026

  Unsaturated compounds, i.e. alkenes, alkynes or allenes · CPC title

• C12Y203/01054

  Formate C-acetyltransferase (2.3.1.54), i.e. pyruvate formate-lyase or PFL · CPC title

• C12P7/649

  Biodiesel, i.e. fatty acid alkyl esters · CPC title