Microbial production of fatty diols

Patent metadata

Field	Value
Publication number	US-10676763-B2
Application number	US-201515326844-A
Country	US
Kind code	B2
Filing date	Jul 17, 2015
Priority date	Jul 18, 2014
Publication date	Jun 9, 2020
Grant date	Jun 9, 2020

Abstract

The disclosure relates to fatty diols and recombinant microorganisms for producing them. More particularly, the disclosure relates to recombinant microorganisms engineered to produce fatty diols via fermentation. Further encompassed is a process that uses the microorganisms to produce fatty diols from a simple carbon source.

First claim

We claim:

1. A recombinant Escherichia coli ( E. coli ) for producing a 1,3 fatty diol when grown in a fermentation broth with a simple carbon source, the E. coli engineered to express heterologous nucleic acid sequences encoding a thioesterase having the classification EC 3.1.2.2, EC 3.1.1.5, or EC 3.1.2.14; and a carboxylic acid reductase having the classification EC 1.2.99.6, wherein the thioesterase uses 3-hydroxy acyl-ACP as a substrate and is selected from the group consisting of TE_EEI82564, TE_CAD63310, and phaG and wherein the carboxylic acid reductase is selected from the group consisting of carB from Mycobacterium smegmatis , carA from Mycobacterium smegmatis , FadD9 from Mycobacterium smegmatis , car from Mycobacterium genavense , car from Nocardia lowensis , and car from Nocardia brasiliensis.

2. The recombinant E. coli of claim 1 , wherein said 1,3 fatty diol is produced in vivo.

3. The recombinant E. coli of claim 1 , wherein said 1,3 fatty diol is selected from the group consisting of a C 5 1,3 fatty diol, a C 6 1,3 fatty diol, a C 7 1,3 fatty diol, a C 8 1,3 fatty diol, a C 9 1,3 fatty diol, a C 10 1,3 fatty diol, a C 11 1,3 fatty diol, a C 12 1,3 fatty diol, a C 13 1,3 fatty diol, a C 14 1,3 fatty diol, a C 15 1,3 fatty diol, a C 16 1,3 fatty diol, a C 17 1,3 fatty diol, a C 18 1,3 fatty diol, and a C 19 1,3 fatty diol.

4. The recombinant E. coli of claim 1 , further expressing a nucleic acid sequence encoding an alcohol dehydrogenase having the classification EC 1.1.1.-.

5. The recombinant E. coli of claim 1 , wherein said simple carbon source is derived from a renewable feedstock.

6. The recombinant E. coli of claim 1 , wherein said carboxylic acid reductase is carB.

7. The recombinant E. coli of claim 4 , wherein said alcohol dehydrogenase is alrA.

8. A cell culture comprising the E. coli according to claim 1 .

9. The cell culture of claim 8 , wherein said cell culture produces 1,3 fatty diols.

10. The cell culture of claim 9 , wherein said 1,3 fatty diol is selected from the group consisting of a C 5 1,3 fatty diol, a C 6 1,3 fatty diol, a C 7 1,3 fatty diol, a C 8 1,3 fatty diol, a C 9 1,3 fatty diol, a C 10 1,3 fatty diol, a C 11 1,3 fatty diol, a C 12 1,3 fatty diol, a C 13 1,3 fatty diol, a C 14 1,3 fatty diol, a C 15 1,3 fatty diol, a C 16 1,3 fatty diol, a C 17 1,3 fatty diol, a C 18 1,3 fatty diol, and a C 19 1,3 fatty diol.

11. A method of producing a 1,3 fatty diol comprising culturing the E. coli of claim 1 .

12. A method of producing a 1,3 fatty diol comprising: (i) culturing a recombinant E. coli in a fermentation broth, wherein said E. coli expresses heterologous nucleic acid sequences encoding a thioesterase having the classification EC 3.1.2.2, EC 3.1.1.5, or EC 3.1.2.14; and a carboxylic acid reductase having the classification EC 1.2.99.6, and optionally an alcohol dehydrogenase having the classification EC 1.1.1.1 or 1.1.1.2; wherein the thioesterase uses 3-hydroxy acyl-ACP as a substrate and is selected from the group consisting of tesA (with leader sequence) from E. coli , tesA (without leader sequence) from E. coli , tes from Photbacterium profundum , tesB from E. coli , TE CAD63310 from Lactobacillus plantarum , TE EEI82564 from Anaerococcus tetraclius , fatB1 from Umbellularia californica , fatB2 from Cuphea hookeriana , fatB3 from Cuphea hookeriana , fatB from Cinnamomurn camphora , fatB from Arabidopsis thaliana , fatA1 from Helianthus animus, fatA from Brassica juncea , fatA from Cuphea hookeriana , fadM from Escherichia coli , yciA from Escherichia coli , ybgC from Escherichia coli , and phaG from Pseudomonas putida , and wherein the carboxylic acid reductase is selected from the group consisting of carB from Mycobacterium smegmatis , carA from Mycobacterium smegmatis , FadD9 from Mycobacterium smegmatis , car from Mycobacterium genavense , car from Nocardia lowensis , and car from Nocardia brasiliensis ; and (ii) isolating a 1,3 fatty diol from said fermentation broth, wherein said fermentation broth comprises a simple carbon source.

13. The method of claim 12 , wherein said 1,3 fatty diol is selected from the group consisting of a C 5 1,3 fatty diol, a C 6 1,3 fatty diol, a C 7 1,3 fatty diol, a C 8 1,3 fatty diol, a C 9 1,3 fatty diol, a C 10 1,3 fatty diol, a C 11 1,3 fatty diol, a C 12 1,3 fatty diol, a C 13 1,3 fatty diol, a C 14 1,3 fatty diol, a C 15 1,3 fatty diol, a C 16 1,3 fatty diol, a C 17 1,3 fatty diol, a C 18 1,3 fatty diol, and a C 19 1,3 fatty diol.

Assignees

• Reg Life Sciences Llc
• Genomatica Inc

Inventors

• Schirmer Andreas W
• Helman Noah
• Wang Haibo
• Hu Zhihao
• Arlagadda Vikranth
• Ramos-Solis Alma Itzel
• Clarke Elizabeth

Classifications

• C12N9/93

  Ligases (6) · CPC title

• C12Y301/02014

  Oleoyl-[acyl-carrier-protein] hydrolase (3.1.2.14), i.e. ACP-thioesterase · CPC title

• C12Y602/01003

  Long-chain-fatty-acid-CoA ligase (6.2.1.3) · CPC title

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title

• C12Y101/01

  with NAD+ or NADP+ as acceptor (1.1.1) · CPC title