# US 2021/0277429 A1 · Muconic acid production from genetically engineered microorganisms

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- Number: US 2021/0277429 A1
- Document ID: US20210277429A1
- Status: Published application
- Kind: A1
- Application number: 17/326,643
- Filed: 2021-05-21
- Published: 2021-09-09
- Earliest claimed: 2016-03-02
- Assignees: PTT Global Chemical Public Company Limited
- Applicants: Ptt Global Chemical Public Company Limited
- Inventors: Ryan SILLERS; Theron HERMANN; Michelle SPENCER; Russell UDANI; R. Rogers YOCUM
- Web page: https://patentgrep.com/patent/US20210277429A1

## CPC

- C12P 7/44: Polycarboxylic acids
- C12P 7/46: Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid
- C12N 9/88: Lyases (4.)
- C12N 15/52: Genes encoding for enzymes or proenzymes
- C12N 15/70: Vectors or expression systems specially adapted for E. coli
- C12Y 401/01063: Protocatechuate decarboxylase (4.1.1.63)
- C12Y 402/01118: 3-Dehydroshikimate dehydratase (4.2.1.118)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES

## Abstract

The subject of this invention is improvements in the yield and titer of biological production of muconic acid by fermentation. Increased activity of one or more enzymes involved in the muconic acid pathway leads to increased production of muconic acid.

## Claims

### Claim at position 1

1-10. (canceled)

### Claim 11

A genetically engineered microorganism that produces muconic acid from a non-aromatic carbon source comprising at least one exogenous gene encoding for 3,4-dihydroxybenzoic acid decarboxylase and ubiX gene from Escherichia coli encoding for UbiX protein that increases the activity of said 3,4-dihydroxybenzoic acid decarboxylase.

### Claim 12 (depends on 11)

The genetically engineered microorganism of claim 11, wherein the genetically engineered microorganism produces at least 60 g/L of muconic acid in 72 hours.

### Claim 13 (depends on 11)

The genetically engineered microorganism of claim 11, wherein the genetically engineered microorganism is a bacterium.

### Claim 14 (depends on 13)

The genetically engineered microorganism of claim 13, wherein the genetically engineered microorganism is Escherichia coli.

### Claim 15 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one catAX exogenous gene encoding for the catechol 1,2-dioxygenase.

### Claim 16 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising one or more exogenous genes encoding for the 3-dehydroshikimate dehydratase selected from aroZ, qa4, asbF and quiC.

### Claim 17 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one aroB exogenous gene encoding for the 3-dehydroquinate synthase.

### Claim 18 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one aroG exogenous gene encoding for the deoxyarabino-heptulosonate.

### Claim 19 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one tktA exogenous gene encoding for the transketolase.

### Claim 20 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one aroD exogenous gene encoding for the 3-dehydroquinate dehydratase.

### Claim 21 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising a mutation or deletion in at least one gene that encode at least one protein that function in a phosphotransferase system for sugar import.

### Claim 22 (depends on 21)

The genetically engineered microorganism of claim 21, wherein the at least one gene is ptsH gene, ptsI gene or a combination thereof.

### Claim 23 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one exogenous gene encoding for a protein that functions in the facilitated diffusion of a sugar.

### Claim 24 (depends on 23)

The genetically engineered microorganism of claim 23, wherein the at least one exogenous gene is a glf gene encoding for the glucose facilitator.

### Claim 25 (depends on 23)

The genetically engineered microorganism of claim 23, further comprising an glk exogenous gene encoding for the glucokinase.

### Claim 26 (depends on 24)

The genetically engineered microorganism of claim 24, further comprising an glk exogenous gene encoding for the glucokinase.

### Claim 27 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising a deletion in a gene encoding for a sugar importer that functions using proton symport.

### Claim 28 (depends on 11)

The genetically engineered microorganism of claim 11, further comprising at least one pyc exogenous gene encoding for the pyruvate carboxylase and a mutation of ppc gene encoding for the phosphoenol pyruvate carboxylase.