Increased production of isobutanol in yeast with reduced mitochondrial amino acid biosynthesis

• US 9,404,117 B2
• Filed: 09/09/2015
• Issued: 08/02/2016
• 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
• Primary Examiner(s): Monshipouri, Maryam
• Application Number:
  US14/848,792
• Publication Number:
  US 20150368656A1
• Time in Patent Office: 328 Days
• Field of Search: 435/252.3
• US Class Current: 1/1
• 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 is substantially devoid of isopropylmalate synthase activity.

2. The recombinant yeast cell of claim 1, wherein the isopropylmalate synthase activity is defined by the enzyme classification number EC 2.3.3.13.

3. The recombinant yeast cell of claim 1, wherein the yeast cell comprises a disruption in a LEU4 gene.

4. The recombinant yeast cell of claim 1, wherein the mitochondria is further substantially devoid of branched chain amino acid transaminase activity.

5. The recombinant yeast cell of claim 4, wherein the branched chain amino acid transaminase activity is defined by the enzyme classification number EC 2.6.1.42.

6. The recombinant yeast cell of claim 4, wherein the yeast cell comprises a disruption in a BAT1 gene.

7. The recombinant yeast cell of claim 1, wherein the mitochondria is further substantially devoid of pyruvate dehydrogenase activity.

8. The recombinant yeast cell of claim 7, wherein the pyruvate dehydrogenase activity is defined by the enzyme classification number EC 1.2.4.1.

9. The recombinant yeast cell of claim 7, wherein the pyruvate dehydrogenase activity is defined by a multienzyme complex comprising proteins selected from the group consisting of PDA1, PDB1, LAT1, LPD1, and PDX1.

10. The recombinant yeast cell of claim 9, wherein the recombinant yeast cell comprises a disruption in a gene encoding a protein selected from the group consisting of PDA1, PDB1, LAT1, LPD1, and PDX1.

11. The recombinant yeast cell of claim 1, wherein the recombinant yeast cell further comprises reduced or eliminated endogenous pyruvate decarboxylase activity.

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