IP Library › Granted Patent US 12,442,026
Granted Patent B2
US 12,442,026 · App. 18/421,170 · Granted Oct 14, 2025

Production of fatty acyl-CoA in yeast using a fatty acid feedstock

Inventors: Alexander Hutagalung (San Diego, CA); Jose Miguel Laplaza (San Diego, CA)
Assignee: Pyrone Systems, Inc.
C12P17/06C12N1/16C12P7/42C12Y103/03006C12Y203/01206C12Y404/01026
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Quick Facts
Patent No.
US 12,442,026
App. No.
18/421,170
Granted
Oct 14, 2025
Kind
B2
Abstract

Strains of yeasts are provided containing the genes for the production of cannabinoids from fatty acids. The enzymes that mediate cannabinoid production are localized to the cytosol, peroxisome or different compartments within the secretory pathway (e.g., endoplasmic reticulum, Golgi, vacuole) to ensure efficient production. The engineered microorganisms produce cannabinoids in a controlled fermentation process.

Claims (22)

1. A method of producing a fatty acyl-CoA in a peroxisome, the method comprising:

providing a fatty acid to a genetically modified microorganism, wherein the genetically modified organism is a yeast or a fungus, wherein the microorganism comprises a biosynthetic enzyme targeted to the peroxisome, wherein the biosynthetic enzyme is capable of catabolizing the fatty acid into the fatty acyl-CoA;

culturing the microorganism under conditions sufficient to produce the fatty acyl-CoA in the peroxisome; and

isolating the fatty acyl-CoA.

2. The method of claim 1 , wherein the fatty acyl-CoA is acetyl-CoA, malonyl-CoA, or hexanoyl-CoA.

3. The method of claim 1 , wherein the yeast is from a genus selected from the group consisting of Candida, Arxula, Pichia, Scheffersomyces, Kluyveromyces, Saccharomyces, Yarrowia , or Schizosaccharomyces.

4. The method of claim 1 , wherein the fungus is Aspergillus parasiticus, Aspergillus nidulans , Thraustochytrium, Schizochytrium, Rhizopus arrhizus, Rhizopus oryzae , or Rhizopus nigricans.

5. The method of claim 1 , wherein the biosynthetic enzyme is a fatty acyl-CoA synthetase, fatty acyl activating enzyme, acyl-CoA oxidase, acyl-CoA thioesterase, hexanoyl-CoA synthetase, acetyl-CoA carboxylase, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, beta-ketothiolase, thiolase, acyl-CoA synthase, cannabidiolic acid synthase, tetrahydrocannabidiolic acid, olivetolic acid synthase, or polyketide synthase.

6. The method of claim 5 , wherein the polyketide synthase is a Type III polyketide synthase.

7. The method of claim 6 , wherein the Type III polyketide synthase is tetraketide synthase, TKS1, or TKS1p.

8. The method of claim 1 , wherein the biosynthetic enzyme comprises a peroxisomal targeting sequence.

9. The method of claim 8 , wherein the peroxisomal targeting sequence has a consensus sequence of [S/A/H/C/E/P/Q/V]-[K/R/H/Q]-[L/F] as set forth in SEQ ID NO: 7, or GRRAKL as set forth in SEQ ID NO: 6.

10. The method of claim 1 , wherein the enzyme is removed of its endogenous amino-terminal localization sequence and/or carboxyl-terminal localization sequence.

11. A genetically modified microorganism, comprising a biosynthetic enzyme targeted to a peroxisome in the microorganism, wherein the genetically modified microorganism is a yeast or a fungus, wherein the biosynthetic enzyme is capable of catabolizing a fatty acid into a fatty acyl-CoA.

12. The genetically modified microorganism of claim 11 , wherein the yeast is from a genus selected from the group consisting of Candida, Arxula, Pichia, Scheffersomyces, Kluyveromyces, Saccharomyces, Yarrowia , or Schizosaccharomyces.

13. The genetically modified microorganism of claim 11 , wherein the fungus is Aspergillus parasiticus, Aspergillus nidulans , Thraustochytrium, Schizochytrium, Rhizopus arrhizus, Rhizopus oryzae , or Rhizopus nigricans.

14. The genetically modified microorganism of claim 11 , wherein the biosynthetic enzyme is a fatty acyl-CoA synthetase, fatty acyl activating enzyme, acyl-CoA oxidase, acyl-CoA thioesterase, hexanoyl-CoA synthetase, acetyl-CoA carboxylase, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, beta-ketothiolase, thiolase, acyl-CoA synthase, cannabidiolic acid synthase, tetrahydrocannabidiolic acid, olivetolic acid synthase, or polyketide synthase.

15. The genetically modified microorganism of claim 11 , wherein the biosynthetic enzyme comprises a peroxisomal targeting sequence.

16. The genetically modified microorganism of claim 15 , wherein the peroxisomal targeting sequence has a consensus sequence of [S/A/H/C/E/P/Q/V]-[K/R/H/Q]-[L/F] as set forth in SEQ ID NO: 7, or GRRAKL as set forth in SEQ ID NO: 6.

17. The genetically modified microorganism of claim 11 , wherein the microorganism is capable of producing in the peroxisome a fatty acyl-CoA.

18. The genetically modified microorganism of claim 17 , wherein the fatty acyl-CoA is acetyl-CoA, malonyl-CoA, or hexanoyl-CoA.

19. The genetically modified microorganism of claim 11 , wherein the enzyme is removed of its endogenous amino-terminal localization sequence and/or carboxyl-terminal localization sequence.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 24, 2024
From: LAPLAZA, JOSE M.; HUTAGALUNG, ALEXANDER
To: LEVADURA BIOTECHNOLOGY, INC.
Reel/Frame 066234/0118 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 24, 2024
From: LEVADURA BIOTECHNOLOGY, INC.
To: PYRONE SYSTEMS, INC.
Reel/Frame 066234/0153 →
Continuity (6)
Continuation 17449847 · Oct 4, 2021
Continuation 16783122 · Feb 5, 2020
Continuation PCTUS2019051357 · Sep 16, 2019
Provisional Application 62731978 · Sep 17, 2018
Provisional Application 62731980 · Sep 17, 2018
Related Publication 20240384314A1 · Nov 21, 2024
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