Methods of producing 7-carbon chemicals from long chain fatty acids via oxidative cleavage

Patent metadata

Field	Value
Publication number	US-9920336-B2
Application number	US-201314139142-A
Country	US
Kind code	B2
Filing date	Dec 23, 2013
Priority date	Dec 31, 2012
Publication date	Mar 20, 2018
Grant date	Mar 20, 2018

Abstract

This document describes biochemical pathways for producing pimelic acid, 7-aminoheptanoic acid, 7-hydroxyheptanoic acid, heptamethylenediamine or 1,7-heptanediol by forming two terminal functional groups, comprised of carboxyl, amine or hydroxyl group, in a C7 aliphatic backbone substrate. These pathways, metabolic engineering and cultivation strategies described herein rely on the fatty acid synthesis pathway and oxidative cleavage of long chain acyl-[acp] intermediates by a monooxgenase (e.g., cytochrome P450) such as that encoded by BioI from microorganisms such as Bacillus subtillis.

First claim

What is claimed is:

1. A method for biosynthesizing 7-aminoheptanoic acid, said method comprising:
(i) providing tetradecanoyl-[acp];
(ii) enzymatically synthesizing pimeloyl-[acp] and heptanoate via (1) oxidative cleavage of tetradecanoyl-[acp] by a monooxgenase classified under EC 1.14.15.12 to form 7-oxoheptanoyl-[acp] and heptanal, and (2) contacting 7-oxoheptanoyl-[acp] and heptanal with an aldehyde dehydrogenase classified under EC 1.2.1.3 or EC 1.2.1.4 to form pimeloyl-[acp] and heptanoate;
(iii) enzymatically converting pimeloyl-[acp] to pimelate semialdehyde via contacting pimeloyl-[acp] with a thioesterase classified under EC 3.1.2.- to form pimelic acid, and contacting pimelic acid with a carboxylate reductase classified under EC 1.2.99.6 to form pimelate semialdehyde, or
enzymatically converting heptanoate to pimelate semialdehyde via contacting heptanoate with a monooxygenase classified under EC 1.14.15.1 or EC 1.14.15.3 to form 7-hydroxyheptanoate, and contacting 7-hydroxyheptanoate with an alcohol dehydrogenase classified under EC 1.1.1.- to form pimelate semialdehyde; and
(iv) enzymatically converting pimelate semialdehyde to 7-aminoheptanoic acid by contacting pimelate semialdehyde with a ω-transaminase classified under EC 2.6.1.-.

2. The method of claim 1 , wherein pimeloyl-[acp] is enzymatically converted to pimelate semialdehyde via contacting pimeloyl-[acp] with a thioesterase classified under EC 3.1.2.- to form pimelic acid, and contacting pimelic acid with a carboxylate reductase classified under EC 1.2.99.6 to form pimelate semialdehyde.

3. The method of claim 1 , wherein heptanoate is enzymatically converted to pimelate semialdehyde via contacting heptanoate with a monooxygenase classified under EC 1.14.15.1 or EC 1.14.15.3 to form 7-hydroxyheptanoate, and contacting 7-hydroxyheptanoate with an alcohol dehydrogenase classified under EC 1.1.1.- to form pimelate semialdehyde.

4. The method of claim 1 , wherein tetradecanoyl-[acp] is provided via a fatty acid synthesis pathway.

5. The method of claim 1 , wherein the monooxgenase classified under EC 1.14.15.12 is encoded by BioI.

6. The method of claim 1 , wherein the thioesterase classified under EC 3.1.2.- has at least 90% sequence identity to the amino acid sequence of SEQ ID NO:1, SEQ ID NO:23, or SEQ ID NO:24.

7. The method of claim 1 , wherein the ω-transaminase classified under EC 2.6.1.- has at least 90% sequence identity to any one of the amino acid sequences of SEQ ID NOs:8-13.

8. The method of claim 1 , wherein the carboxylate reductase classified under EC 1.2.99.6 has at least 90% sequence identity to any one of the amino acid sequences of SEQ ID NOs:2-7.

9. The method of claim 1 , wherein the alcohol dehydrogenase classified under EC 1.1.1.- is a 6-hydroxyhexanoate dehydrogenase classified under EC 1.1.1.258, a 5-hydroxypentanoate dehydrogenase classified under EC 1.1.1.-, or a 4-hydroxybutyrate dehydrogenase classified under EC 1.1.1.-.

10. The method of claim 1 , wherein the ω-transaminase classified under EC 2.6.1.- is a ω-transaminase classified under EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82.

11. The method of claim 1 , wherein said method is performed in a recombinant host by fermentation.

12. The method of claim 11 , wherein the principal carbon source fed to the fermentation derives from biological or non-biological feedstocks.

13. The method of claim 12 , wherein the biological feedstock is, or derives from, monosaccharides, disaccharides, lignocellulose, hemicellulose, cellulose, lignin, levulinic acid, formic acid, triglycerides, glycerol, fatty acids, agricultural waste, condensed distillers' solubles, or municipal waste; and wherein the non-biological feedstock is, or derives from, natural gas, syngas, CO 2 /H 2 , methanol, ethanol, benzoate, non-volatile residue (NVR) caustic wash waste stream from cyclohexane oxidation processes, or terephthalic acid/isophthalic acid mixture waste streams.

14. The method of claim 11 , wherein the recombinant host is a prokaryote.

15. The method of claim 14 , wherein said prokaryote is selected from Escherichia, Clostridia, Corynebacteria, Cupriavidus, Pseudomonas, Delftia, Bacillus, Lactobacillus, Lactococcus , and Rhodococcus.

16. The method of 14 , wherein said prokaryote is selected from Escherichia coli, Clostridium ljungdahlii, Clostridium autoethanogenum, Clostridium kluyveri, Corynebacterium glutamicum, Cupriavidus necator, Cupriavidus metallidurans, Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas oleavorans, Delftia acidovorans, Bacillus subtilis, Lactobacillus delbrueckii, Lactococcus lactis , and Rhodococcus equi.

17. The method of claim 11 , wherein the recombinant host is a eukaryote.

18. The method of claim 17 , wherein said eukaryote is selected from Aspergillus, Saccharomyces, Pichia, Yarrowia, Issatchenkia, Debaryomyces, Arxula , and Kluyveromyces.

19. The method claim 17 , wherein said eukaryote is selected from Aspergillus niger, Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, Issathenkia orientalis, Debaryomyces hansenii, Arxula adenoinivorans , and Kluyveromyces lactis.

20. The method of claim 1 , wherein the monooxygenase classified under EC 1.14.15.1 or EC 1.14.15.3 has at least 90% sequence identity to any one of the amino acid sequences of SEQ ID NOs:14-16.

Assignees

• Invista North America Sarl

Inventors

• Pearlman Paul S
• Conradie Alex Van Eck
• Chen Changlin
• Botes Adriana Leonora

Classifications

• C12P7/46 Primary

  Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid · CPC title

• C12P7/18 Primary

  polyhydric · CPC title

• C12P7/44

  Polycarboxylic acids · CPC title

• C12P13/001

  Amines; Imines · CPC title

• C12P13/005

  Amino acids other than alpha- or beta amino acids, e.g. gamma amino acids · CPC title