Microbial Production of 2-Phenylethanol from Renewable Substrates

Patent metadata

Field	Value
Publication number	US-2020231992-A1
Application number	US-201816633525-A
Country	US
Kind code	A1
Filing date	Jul 18, 2018
Priority date	Jul 25, 2017
Publication date	Jul 23, 2020
Grant date	—

Abstract

Described herein are engineered metabolic pathways in recombinant microorganism host cells which result in the production of 2-phenylethanol or 2-phenylacetic acid. Also described herein are methods of using the recombinant microorganisms for the production of 2-phenylethanol or 2-phenylacetic acid.

First claim

We claim:

1 . A recombinant organism comprising,
(i) at least one heterologous gene encoding an enzyme having phenylalanine ammonia lyase (PAL) activity, (ii) at least one heterologous gene encoding an enzyme having trans-cinnamic acid decarboxylase (CADC) activity, (iii) at least one heterologous gene encoding an enzyme having styrene monooxygenase (SMO) activity, (iv) at least one heterologous gene encoding an enzyme having styrene oxide isomerase (SOI) activity, and (v) at least one gene encoding an enzyme having 2-phenylacetaldehyde reductase (PAR) activity, wherein the recombinant microorganism is capable of producing 2-phenylethanol from a fermentable carbon substrate.

2 . The organism of claim 1 , wherein the organism is Escherichia coli.

3 . The organism of claim 2 , wherein the organism is a phenylalanine overproducing strain of E. coli.

4 . The organism of claim 1 , wherein the gene encoding a polypeptide having phenylalanine ammonia lyase activity is derived from Arabidopsis thaliana.

5 . The organism of claim 1 , wherein the gene encoding polypeptides having trans-cinnamic acid decarboxylase activity is derived from Saccharomyces cerevisiae.

6 . The organism of claim 1 , wherein the gene encoding a polypeptide having styrene monooxygenase activity is derived from Pseudomonas putida.

7 . The organism of claim 1 , wherein the gene encoding a polypeptide having styrene oxide isomerase activity is derived from Pseudomonas putida.

8 . A method of producing 2-phenylethanol comprising the steps of,
(i) contacting the recombinant organism of claim 1 with a fermentable carbon substrate, and (ii) growing the recombinant organism for a time sufficient to produce 2-phenylethanol.

9 . The method of claim 8 , wherein the fermentable carbon substrate is selected from the group consisting of monosaccharides, oligosaccharides, polysaccharides, glycerol, carbon dioxide, methanol, methane, formaldehyde, formate, amino acids, carbon-containing amines.

10 . The method of claim 8 , wherein the fermentable carbon substrate is selected from the group consisting of glucose, xylose, or glycerol.

11 . The method of claim 8 , wherein the fermentable carbon substrate is selected from the group consisting of lignin-derived aromatic monomers, lignin-derived aromatic oligomers and combinations thereof.

12 . The method of claim 8 , wherein the fermentable carbon substrate is a biomass hydrolysate.

13 . A recombinant organism comprising,
(i) at least one heterologous gene encoding an enzyme having phenylalanine ammonia lyase (PAL) activity, (ii) at least one heterologous gene encoding an enzyme having trans-cinnamic acid decarboxylase (CADC) activity, (iii) at least one heterologous gene encoding an enzyme having styrene monooxygenase (SMO) activity, (iv) at least one heterologous gene encoding an enzyme having styrene oxide isomerase (SOI) activity, and (v) at least one gene encoding an enzyme having 2-phenylacetaldehyde dehydrogenase (PADH) activity, wherein the engineered microorganism is capable of producing 2-phenylacetic acid from a fermentable carbon substrate.

14 . The organism of claim 13 , wherein the organism is Escherichia coli.

15 . The organism of claim 14 , wherein the organism is a phenylalanine overproducing strain of E. coli.

16 . The organism of claim 13 , wherein the gene encoding a polypeptide having phenylalanine ammonia lyase activity is derived from Arabidopsis thaliana.

17 . The organism of claim 13 , wherein the gene encoding polypeptides having trans-cinnamic acid decarboxylase activity is derived from Saccharomyces cerevisiae.

18 . The organism of claim 13 , wherein the gene encoding a polypeptide having styrene monooxygenase activity is derived from Pseudomonas putida.

19 . The organism of claim 13 , wherein the gene encoding a polypeptide having styrene oxide isomerase activity is derived from Pseudomonas putida.

20 . A method of producing 2-phenylacetic acid comprising the steps of,
(i) contacting the recombinant organism of claim 13 with a fermentable carbon source, and (ii) growing the recombinant organism for a time sufficient to produce 2-phenylacetic acid.

21 . The method of claim 20 , wherein the fermentable carbon source is selected from the group consisting of monosaccharides, oligosaccharides, polysaccharides, glycerol, carbon dioxide, methanol, methane, formaldehyde, formate, amino acids, carbon-containing amines.

22 . The method of claim 20 , wherein the fermentable carbon source is selected from the group consisting of glucose, xylose, or glycerol.

23 . The method of claim 20 , wherein the fermentable carbon substrate is selected from the group consisting of lignin-derived aromatic monomers, lignin-derived aromatic oligomers, and combinations thereof.

24 . The method of claim 20 , wherein the fermentable carbon substrate is a biomass hydrolysate.

Assignees

• Univ Arizona State

Inventors

• Nielsen David
• Machas Michael
• Mckenna Rebekah

Classifications

• C12R2001/19

  Escherichia coli · CPC title

• C12N1/205

  Bacterial isolates · CPC title

• C12N9/88 Primary

  Lyases (4.) · CPC title

• C12Y403/01024

  Phenylalanine ammonia-lyase (4.3.1.24) · CPC title

• C12Y102/01039

  Phenylacetaldehyde dehydrogenase (1.2.1.39) · CPC title