Microorganisms Engineered to Produce Phenol and Its Derivatives

Patent metadata

Field	Value
Publication number	US-2016160244-A1
Application number	US-201414908967-A
Country	US
Kind code	A1
Filing date	Aug 12, 2014
Priority date	Aug 27, 2013
Publication date	Jun 9, 2016
Grant date	—

Abstract

Novel methods for the in vivo production of phenol from renewable substrates using a recombinant microorganism (FIG. 1 ). Additionally, methods for the in vivo production of catechol and cis,cis-muconic acid from renewable substrates using a recombinant microorganism are disclosed. A host cell expresses at least one gene encoding a polypeptide that possesses isochorismate synthase activity, at least one gene encoding a polypeptide that possesses isochorismate pyruvate lyase activity, and at least one gene encoding a polypeptide that possesses salicylic acid decarboxylase activity. In the case of catechol, the host cell must additionally express at least one gene encoding a polypeptide that possesses phenol 2-monooxy-genase activity. In the case of cis,cis-muconic acid, the host cell must additionally express at least one gene encoding a polypeptide that possesses phenol 2-monooxygenase activity and at least one gene encoding a polypeptide that possesses catechol-1,2-dioxygenase activity.

First claim

1 . A method for the production of phenol comprising:
(i) contacting a recombinant host cell with a fermentable carbon substrate, said recombinant host cell comprising: a) at least one gene encoding a polypeptide having isochorismate synthase activity; b) at least one gene encoding a polypeptide having isochorismate pyruvate lyase activity; and c) at least one gene encoding a polypeptide having salicylate decarboxylase activity; (ii) growing said recombinant cell for a time sufficient to produce phenol.

2 . A method for the production of catechol comprising:
(i) contacting a recombinant host cell with a fermentable carbon substrate, said recombinant host cell comprising: a) at least one gene encoding a polypeptide having isochorismate synthase activity; b) at least one gene encoding a polypeptide having isochorismate pyruvate lyase activity; c) at least one gene encoding a polypeptide having salicylate decarboxylase activity; and d) at least one gene encoding a polypeptide having phenol 2-monooxygenase activity; (ii) growing said recombinant cell for a time sufficient to produce catechol.

3 . A method for the production of cis,cis-muconate comprising:
(i) contacting a recombinant host cell with a fermentable carbon substrate, said recombinant host cell comprising: a) at least one gene encoding a polypeptide having isochorismate synthase activity; b) at least one gene encoding a polypeptide having isochorismate pyruvate lyase activity; c) at least one gene encoding a polypeptide having salicylate decarboxylase activity; d) at least one gene encoding a polypeptide having phenol 2-monooxygenase activity; and e) at least one gene encoding a polypeptide having catechol-1,2-dioxygenase activity; (ii) growing said recombinant cell for a time sufficient to produce catechol.

4 . A method according to claim 1 , wherein the sequence of gene encoding a polypeptide having isochorismate synthase activity is as set forth in SEQ ID NO:1, 2, 3 or 4.

5 . A method according to claim 1 , wherein the sequence of gene encoding a polypeptide having isochorismate pyruvate lyase activity is as set forth in SEQ ID NO:5.

6 . A method according to claim 1 , wherein the sequence of gene encoding a polypeptide having salicylate decarboxylase activity is as set forth in SEQ ID NO:6 or 7.

7 . A method according to claim 2 , wherein the sequence of gene encoding a polypeptide having phenol 2-monooxygenase activity.

8 . A method according to claim 3 wherein the sequence of gene encoding a polypeptide having catechol-1,2-dioxygenase activity.

9 . A method according to claim 1 , wherein said fermentable carbon source is selected from the group consisting of monosaccharides, oligosaccharides, polysaccharides, glycerol, carbon dioxide, methanol, formaldehyde, formate, amino acids, and carbon-containing amines.

10 . A method according to claim 9 wherein said fermentable carbon source is selected from the group consisting of glucose or glycerol.

11 . A method according to claim 1 , wherein said recombinant host cell is selected from the group consisting of bacteria, yeast, filamentous fungi, cyanobacteria, algae, and plant cells.

12 . A method according to claim 11 wherein said recombinant host cell is selected from the group consisting of Escherichia, Salmonella, Bacillus, Acinetobacter, Streptomyces, Corynebacterium, Methylosinus, Methylomonas, Rhodococcus, Pseudomonas, Rhodobacter, Synechocystis, Saccharomyces, Klebsiella, Zygosaccharomyces, Kluyveromyces, Candida, Hansenula, Debaryomyces, Mucor, Pichia, Torulopsis, Aspergillus, Arthrobotrys, Brevibacterium, Microbacterium, Arthrobacter, Ctirobacter, Chlamydomonas , and Zymomonas.

13 . A method according to claim 11 wherein said recombinant host cell is strain that overproduces chorismate or aromatic amino acids.

14 . A method according to claim 1 , wherein said recombinant host cell is a cell isolated from a plant selected from the group consisting of soybean, rapeseed, sunflower, cotton, corn, tobacco, alfalfa, wheat, barley, oats, sorghum, rice, broccoli, cauliflower, cabbage, parsnips, melons, carrots, grapes, grass seed crops, sugar beets, sugar cane, beans, peas, rye, flax, hardwood trees, softwood trees, and forage grasses.

15 . A method according to claim 1 , wherein the gene encoding a polypeptide having isochorismate synthase activity is derived from A. thaliana.

16 . A method according to claim 1 , wherein the gene encoding a polypeptide having isochrosimate synthase activity is derived from P. aeruginosa.

17 . A method according to claim 1 , wherein the gene encoding a polypeptide having isochrosimate synthase activity is derived from E. coli.

18 . A method according to claim 1 , wherein the genes encoding polypeptides having isochorismate pyruvate lyase activity are derived from P. aeruginosa.

19 . A method according to claim 1 wherein the genes encoding polypeptides having salicylate decarboxylase activity are derived from T. moniliiforme.

20 . A method according to claim 3 wherein the gene encoding a polypeptide having phenol 2-monooxygenase activity is derived from T. cutaneum.

21 . A method according to claim 3 wherein the gene encoding a polypeptide having phenol 2-monooxygenase activity is derived from Pseudomonas sp. CF600.

22 . A method according to claim 3 wherein the gene encoding a polypeptide having catechol-1,2-dioxygenase activity is derived from Pseudomonas reinekei.

23 . A method according to claim 3 wherein the gene encoding a polypeptide having catechol-1,2-dioxygenase activity is derived from Pseudomonas putida.

Assignees

• Nielsen David
• Pugh Shawn
• Thompson Brian
• Univ Arizona State

Inventors

• Nielsen David
• Pugh Shawn
• Thompson Brian

Classifications

• C12N9/90

  Isomerases (5.) · CPC title

• C12N9/88

  Lyases (4.) · CPC title

• C12P7/44

  Polycarboxylic acids · CPC title

• C12P7/22 Primary

  aromatic · CPC title