Method for the in vivo synthesis of 4-hydroxymethylfurfural and derivatives thereof

Patent metadata

Field	Value
Publication number	US-11566269-B2
Application number	US-202117163838-A
Country	US
Kind code	B2
Filing date	Feb 1, 2021
Priority date	Mar 1, 2019
Publication date	Jan 31, 2023
Grant date	Jan 31, 2023

Abstract

The present disclosure provides recombinant microorganisms and methods for the production of 4-HMF, 2,4-furandimethanol, furan-2,4-dicarbaldehyde, 4-(hydroxymethyl)furoic acid, 2-formylfuran-4-carboxylate, 4-formylfuran-2-carboxylate, and/or 2,4-FDCA from a carbon source. The method provides for engineered microorganisms that express endogenous and/or exogenous nucleic acid molecules that catalyze the conversion of a carbon source into 4-HMF, 2,4-furandimethanol, furan-2,4-dicarbaldehyde, 4-(hydroxymethyl)furoic acid, 2-formylfuran-4-carboxylate, 4-formylfuran-2-carboxylate, and/or 2,4-FDCA. The disclosure further provides methods of producing polymers derived from 4-HMF, 2,4-furandimethanol, furan-2,4-dicarbaldehyde, 4-(hydroxymethyl)furoic acid, 2-formylfuran-4-carboxylate, 4-formylfuran-2-carboxylate, and/or 2,4-FDCA.

First claim

What is claimed:

1. A recombinant microorganism capable of producing 2,4-furandimethanol (2,4-FDME) from a feedstock comprising a carbon source, wherein the recombinant microorganism expresses the following:
(a) endogenous or exogenous nucleic acid molecules encoding enzymes capable of converting a carbon source to glyceraldehyde 3-phosphate (G3P);
(b) at least one exogenous nucleic acid molecule encoding a (5-formylfuran-3-yl)methyl phosphate synthase that catalyzes the conversion of G3P from (a) to (5-formylfuran-3-yl)methyl phosphate;
(c) at least one endogenous or exogenous nucleic acid molecule encoding a phosphatase that catalyzes the conversion of (5-formylfuran-3-yl)methyl phosphate from (b) to 4-hydroxymethylfurfural (4-HMF); and
(d) at least one endogenous or exogenous nucleic acid molecule encoding a dehydrogenase that catalyzes the conversion of 4-HMF from (c) to 2,4-FDME.

2. The recombinant microorganism of claim 1 , wherein the carbon source comprises a hexose, a pentose, glycerol, CO 2 , sucroses or combinations thereof.

3. The recombinant microorganism of claim 1 , wherein the (5-formylfuran-3-yl)methyl phosphate synthase comprises an amino acid sequence comprising SEQ ID NO: 1, SEQ ID NO: 7, or SEQ ID NO: 14.

4. The recombinant microorganism of claim 1 , wherein the phosphatase is endogenous to the recombinant microorganism.

5. The recombinant microorganism of claim 4 , wherein phosphatase endogenous to the recombinant microorganism is overexpressed.

6. The recombinant microorganism of claim 1 , wherein the phosphatase is encoded by an amino acid sequence comprising SEQ ID NO: 28, any one of SEQ ID NOs 40-52, or any one of SEQ ID NOs 53-68.

7. The recombinant microorganism of claim 1 , wherein the dehydrogenase is an alcohol dehydrogenase.

8. The recombinant microorganism of claim 7 , wherein the dehydrogenase is endogenous to the recombinant microorganism.

9. The recombinant microorganism of claim 8 , wherein the dehydrogenase endogenous to the recombinant microorganism is overexpressed.

10. The recombinant microorganism of claim 7 , wherein the dehydrogenase is an alcohol dehydrogenase NADPH.

11. The recombinant microorganism of claim 7 , wherein the dehydrogenase is an alcohol dehydrogenase NADH.

12. The recombinant microorganism of claim 7 , wherein the dehydrogenase comprises an amino acid sequence comprising SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, or SEQ ID NO: 92.

13. The recombinant microorganism of claim 1 , wherein the one or more recombinant microorganisms are derived from a parental microorganism selected from the group consisting of Clostridium sp., Clostridium ljungdahlii, Clostridium autoethanogenum, Clostridium ragsdalei, Eubacterium limosum, Butyribacterium methylotrophicum, Moorella thermoacetica, Corynebacterium glutamicum, Clostridium aceticum, Acetobacterium woodii, Alkalibaculum bacchii, Clostridium drakei, Clostridium carboxidivorans, Clostridium formicoaceticum, Clostridium scatologenes, Moorella thermoautotrophica, Acetonema longum, Blautia producta, Clostridium glycolicum, Clostridium magnum, Candida krusei, Clostridium mayombei, Clostridium methoxybenzovorans, Clostridium acetobutylicum, Clostridium beijerinckii, Oxobacter pfennigii, Thermoanaerobacter kivui, Sporomusa ovata, Thermoacetogenium phaeum, Acetobacterium carbinolicum, Issatchenkia orientalis, Sporomusa termitida, Moorella glycerini, Eubacterium aggregans, Treponema azotonutricium, Pichia kudriavzevii, Escherichia coli, Saccharomyces cerevisiae, Pseudomonas putida, Bacillus sp, Corynebacterium sp., Yarrowia lipolytica, Scheffersomyces stipitis , and Terrisporobacter glycolicus.

14. A method of producing 2,4-FDME using a recombinant microorganism of claim 1 , the method comprising cultivating the recombinant microorganism in a culture medium containing a feedstock providing a carbon source until the 2,4-FDME is produced.

Assignees

• Braskem Sa

Inventors

• Alexandrino Paulo Moises Raduan
• Gouvea Iuri Estrada
• Queiroz Veronica Leite
• Simões Marcos Rogerio

Classifications

• C12P17/04 Primary

  containing a five-membered hetero ring, e.g. griseofulvin {, vitamin C} · CPC title

• C12Y102/01

  with NAD+ or NADP+ as acceptor (1.2.1) · CPC title

• C12Y402/03

  acting on phosphates (4.2.3) · CPC title

• C12Y301/03

  Phosphoric monoester hydrolases (3.1.3) · CPC title

• C12Y102/01005

  Aldehyde dehydrogenase [NAD(P)+] (1.2.1.5) · CPC title