INCREASED PRODUCTION OF ISOBUTANOL IN YEAST WITH REDUCED MITOCHONDRIAL AMINO ACID BIOSYNTHESIS

• US 20100129887A1
• Filed: 11/12/2009
• Published: 05/27/2010
• Est. Priority Date: 11/13/2008
• Status: Active Grant

• Current Assignee: Gevo Incorporated
• Original Assignee: Butamax(TM) Advanced Biofuels LLC (BP PLC)
• Inventors: ANTHONY, Larry Cameron
• Granted Patent: US 8,465,964 B2
• Time in Patent Office: Days
• US Class Current: 435/160
• CPC Class Codes: C12N 15/81 for yeasts
• CPC Class Codes: C12N 9/1025 Acyltransferases (2.3)
• CPC Class Codes: C12N 9/1029 transferring groups other than amino-acyl groups (2.3.1)
• CPC Class Codes: C12N 9/1096 transferring nitrogenous groups (2.6)
• CPC Class Codes: C12N 9/88 Lyases (4.)
• CPC Class Codes: C12P 7/16 Butanols
• CPC Class Codes: C12Y 203/03013 2-Isopropylmalate synthase (2.3.3.13)
• CPC Class Codes: C12Y 206/01042 Branched-chain-amino-acid transaminase (2.6.1.42)
• CPC Class Codes: C12Y 401/01001 Pyruvate decarboxylase (4.1.1.1)
• CPC Class Codes: C12Y 403/01019 Threonine ammonia-lyase (4.3.1.19)
• CPC Class Codes: Y02E 50/10 Biofuels, e.g. bio-diesel
• CPC Class Codes: Y02P 20/52 using catalysts, e.g. selective catalysts

Abstract

Yeast cells with reduced activity of certain enzymes involved in branched chain amino acid biosynthesis in yeast mitochondria are described. Target enzymes include threonine deaminase, isopropylmalate synthase, and optionally branched chain amino acid transaminase.

Claims

1. A recombinant yeast host cell which produces isobutanol and comprises mitochondria which are substantially devoid of an enzyme activity selected from the group consisting of threonine deaminase and isopropylmalate synthase activity.

2. The yeast cell of claim 1 wherein the mitochondria is substantially devoid of branched chain amino acid transaminase activity.

3. The yeast cell of claim 1 wherein the mitochondria is substantially devoid of pyruvate dehydrogenase activity.

4. The yeast cell of claim 1 wherein the threonine deaminase activity is defined by the enzyme classification number EC 4.3.1.19 and the isopropylmalate synthase activity is defined by the enzyme classification number EC 2.3.3.13.

5. The yeast cell of claim 1 wherein endogenous pyruvate decarboxylase activity is reduced.

6. The yeast cell of claim 2 wherein the branched chain amino acid transaminase activity is defined by the enzyme classification number EC 2.6.1.42.

7. The yeast cell of claim 3 pyruvate dehydrogenase activity is defined by the enzyme classification number EC 1.2.4.1.

8. The yeast cell of claim 3 wherein the pyruvate dehydrogenase activity is defined by a multienzyme complex comprising proteins selected from the group consisting of:

PDA1, PDA1, PDB1, LAT1, LPD1, and PDX1.

9. The recombinant yeast host cell of claim 1 wherein the yeast is selected from the group consisting of Saccharomyces, Schizosaccharomyces, Hansenula, Candida, Kluyveromyces, Yarrowia and Pichia.

10. The yeast cell of claim 1 comprising a disruption in a gene selected from the group consisting of ILV1 and LEU4.

11. The yeast cell of claim 2 comprising a disruption in the BAT1 gene.

12. The yeast cell of claim 3 comprising a disruption in a gene encoding a protein selected from the group consisting of PDA1, PDA1, PDB1, LAT1, LPD1, and PDX1

13. A method for the production of isobutanol comprising growing the host cell of claim 1 under conditions wherein isobutanol is produced.

14. A method for the production of isobutanol comprising growing the host cell of claim 2 under conditions wherein isobutanol is produced.

15. A method for the production of isobutanol comprising growing the host cell of claim 3 under conditions wherein isobutanol is produced.