Production of carotenoids in oleaginous yeast and fungi

• US 9,909,130 B2
• Filed: 08/15/2012
• Issued: 03/06/2018
• Est. Priority Date: 03/18/2005
• Status: Active Grant

• Current Assignee: DSM IP Assets BV (DSM-Firmenich AG)
• Original Assignee: DSM IP Assets BV (DSM-Firmenich AG)
• Inventors: Bailey, Richard B., Madden, Kevin T., Trueheart, Joshua
• Primary Examiner(s): Ketter, Jim
• Application Number:
  US13/586,611
• Publication Number:
  US 20130045504A1
• Time in Patent Office: 2,029 Days
• Field of Search: None
• CPC Class Codes: A23L 31/00 Edible extracts or preparations of fungi; Preparation or treatment thereof
• CPC Class Codes: A23L 31/10 Yeasts or derivatives thereof
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/04 specific for metastasis
• CPC Class Codes: A61P 37/08 Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 9/04 Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: A61P 9/12 Antihypertensives
• CPC Class Codes: C12N 15/80 for fungi
• CPC Class Codes: C12P 23/00 : Preparation of compounds containing a cyclohexene ring having an unsaturated side chain containing at least ten carbon atoms bound by conjugated double bonds, e.g. carotenes containing heterorings C12P17/00

Abstract

The present invention provides systems for producing engineered oleaginous yeast or fungi that express carotenoids.

Claims

1. A recombinant oleaginous fungus selected from Yarrowia or Saccharomyces characterized by:

i. lipid accumulation to at least 20% of its dry weight; and

ii. as a result of a recombinant process, production of at least one carotenoid and accumulation of the at least one carotenoid to at least 1% of its dry cell weight, wherein the at least one carotenoid is selected from the group consisting of beta-carotene and astaxanthin.

2. The recombinant fungus of claim 1, wherein the lipid accumulation to at least 20% of its dry cell weight occurs as a result of genetic engineering.

3. The recombinant fungus of claim 2, wherein the fungus is selected from a species group consisting of Yarrowia lipolytica and Saccharomyces cerevisiae.

4. The recombinant fungus of claim 1, wherein the at least one carotenoid is accumulated in cytoplasmic oil bodies.

5. The recombinant fungus of claim 1, wherein the recombinant process alters the expression or activity of a carotenogenic polypeptide selected from the group consisting of isoprenoid biosynthesis polypeptides, carotenoid biosynthesis polypeptides, isoprenoid biosynthesis competitor polypeptides, and combinations thereof.

6. The recombinant fungus of claim 5, wherein the carotenoid biosynthesis polypeptide is selected from the group consisting of:

phytoene synthase polypeptide, phytoene dehydrogenase polypeptide, lycopene cyclase polypeptide, carotenoid ketolase polypeptide, carotenoid hydroxylase polypeptide, astaxanthin synthase polypeptide, carotenoid epsilon hydroxylase polypeptide, carotenoid glucosyltransferase polypeptide, lycopene cyclase (beta and epsilon subunits) polypeptides, and acyl CoA;

diacyglycerol acyltransferase polypeptide.

7. The genetically recombinant fungus of claim 5, wherein the carotenogenic polypeptide confers to the yeast the ability to produce the at least one carotenoid to a level at least 1% of its dry cell weight or confers to the yeast the ability to produce at least one carotenoid which the yeast does not naturally produce.

8. The recombinant fungus of claim 5, wherein the fungus comprises at least one modification selected from the group consisting of:

i. expression of truncated HMG-CoA reductase;

ii. heterologous expression of carotene hydroxylase and carotene ketolase;

iii. complete or partial inactivation of a gene encoding squalene synthase; and

iv. combinations thereof.

9. The recombinant fungus of claim 1, further characterized by an oleaginic modification.

10. The recombinant fungus of claim 9, wherein the oleaginic modification confers upon the fungus an ability to accumulate lipid to at least 20% of its dry cell weight.

11. The recombinant fungus of claim 9, wherein the oleaginic modification alters expression or activity of at least one oleaginic polypeptide.

12. The recombinant fungus of claim 11, wherein the at least one oleaginic polypeptide is selected from the group consisting of acetyl-CoA carboxylase polypeptide, pyruvate decarboxylase polypeptide, isocitrate dehydrogenase polypeptide, ATP citrate lyase polypeptide, malic enzyme polypeptide, AMP deaminase polypeptide, and combinations thereof.