Isobutanol production using yeasts with modified transporter expression

Patent metadata

Field	Value
Publication number	US-2016108441-A1
Application number	US-201514976563-A
Country	US
Kind code	A1
Filing date	Dec 21, 2015
Priority date	May 31, 2010
Publication date	Apr 21, 2016
Grant date	—

Abstract

The present invention relates to recombinant microorganisms comprising an isobutanol producing metabolic pathway and methods of using said recombinant microorganisms to produce isobutanol. In various aspects of the invention, the recombinant microorganisms may comprise one or more modifications resulting in the reduction in the expression and/or activity of an endogenous transporter protein. In various embodiments described herein, the recombinant microorganisms may be microorganisms of the Saccharomyces clade, Crabtree-negative yeast microorganisms, Crabtree-positive yeast microorganisms, post-WGD (whole genome duplication) yeast microorganisms, pre-WGD (whole genome duplication) yeast microorganisms, and non-fermenting yeast microorganisms.

First claim

1 - 20 . (canceled)

21 . A recombinant yeast microorganism comprising enzymes of an isobutanol producing metabolic pathway, comprising at least one exogenous gene encoding a polypeptide that catalyzes a step in the conversion of pyruvate to isobutanol, wherein said recombinant microorganism is engineered to reduce the expression and/or activity of one or more of the endogenous transporter proteins encoded by a gene selected from AQR1 and PDR12.

22 . The recombinant yeast microorganism of claim 21 , wherein the expression and/or activity is reduced by inactivating mutations, disruption, partial deletion, or completely deletion of the endogenous transporter proteins.

23 . The recombinant yeast microorganism of claim 21 , wherein said recombinant microorganism is engineered to disrupt, partially delete, or completely delete, or inactivate via one or more point mutations, a gene encoding pyruvate decarboxylase.

24 . The recombinant yeast microorganism of claim 21 , wherein said recombinant microorganism is engineered to disrupt, partially delete, or completely delete, or inactivate via one or more point mutations, a gene encoding glycerol-3-phosphate dehydrogenase.

25 . The recombinant yeast microorganism of claim 21 , wherein said recombinant microorganism is of the Saccharomyces clade.

26 . The recombinant yeast microorganism of claim 25 , wherein said recombinant microorganism is a Saccharomyces sensu stricto microorganism.

27 . The recombinant yeast microorganism of claim 26 , wherein said Saccharomyces sensu stricto microorganism is selected from the group consisting of S. cerevisiae, S. kudriavzevii, S. mikatae, S. bayanus, S. uvarum, S. carocanis and hybrids thereof.

28 . The recombinant yeast microorganism of claim 21 , wherein said recombinant microorganism is a Crabtree-negative yeast microorganism.

29 . The recombinant yeast microorganism of claim 28 , wherein said Crabtree-negative yeast microorganism is classified into a genus selected from a group consisting of Saccharomyces, Kluyveromyces, Pichia, Hansenula, Issatchenkia and Candida.

30 . The recombinant yeast microorganism of claim 29 , wherein said Crabtree-negative yeast microorganism is selected from the group consisting of Saccharomyces kluyveri, Kluyveromyces lactis, Kluyveromyces marxianus, Pichia anomala, Pichia stipitis, Pichia kudriavzevii, Issatchenkia orientalis, Hansenula anomala, Candida utilis and Kluyveromyces waltii.

31 . The recombinant yeast microorganism of claim 21 , wherein said recombinant microorganism is a Crabtree-positive yeast microorganism.

32 . The recombinant yeast microorganism of claim 31 , wherein said Crabtree-positive yeast microorganism is classified into a genus selected from a group consisting of Saccharomyces, Kluyveromyces, Zygosaccharomyces, Debaryomyces, Pichia, Candida , and Schizosaccharomyces.

33 . The recombinant microorganism of claim 32 , wherein said Crabtree-positive yeast microorganism is selected from the group consisting of Saccharomyces cerevisiae, Saccharomyces uvarum, Saccharomyces bayanus, Saccharomyces paradoxus, Saccharomyces castelli, Kluyveromyces thermotolerans, Candida glabrata, Zygosaccharomyces bailli, Zygosaccharomyces rouxii, Debaryomyces hansenii, Pichia pastorius, Schizosaccharomyces pombe , and Saccharomyces uvarum.

34 . The recombinant yeast microorganism of claim 21 , wherein said recombinant yeast microorganism is a post-WGD (whole genome duplication) yeast microorganism.

35 . The recombinant yeast microorganism of claim 23 , wherein said post-WGD yeast microorganism is classified into a genus selected from a group consisting of Saccharomyces or Candida.

36 . The recombinant yeast microorganism of claim 24 , wherein said post-WGD yeast microorganism is selected from the group consisting of Saccharomyces cerevisiae, Saccharomyces uvarum, Saccharomyces bayanus, Saccharomyces paradoxus, Saccharomyces castelli , and Candida glabrata.

37 . The recombinant yeast microorganism of claim 21 , wherein said recombinant yeast microorganism is a pre-WGD (whole genome duplication) yeast microorganism.

38 . The recombinant yeast microorganism of claim 26 , wherein said pre-WGD yeast microorganism is classified into a genus selected from a group consisting of Saccharomyces, Kluyveromyces, Candida, Pichia, Debaryomyces, Hansenula, Issatchenkia, Pachysolen, Yarrowia and Schizosaccharomyces.

39 . The recombinant yeast microorganism of claim 27 , wherein said pre-WGD yeast microorganism is selected from the group consisting of Saccharomyces kluyveri, Kluyveromyces thermotolerans, Kluyveromyces marxianus, Kluyveromyces waltii, Kluyveromyces lactis, Candida tropicalis, Pichia pastoris, Pichia anomala, Pichia stipitis, Pichia kudriavzevii, Issatchenkia orientalis, Debaryomyces hansenii, Hansenula anomala, Pachysolen tannophilis, Yarrowia lipolytica , and Schizosaccharomyces pombe.

40 . A method of producing isobutanol, comprising:
(a) providing a recombinant microorganism of claim 21 ; (b) cultivating the recombinant microorganism in a culture medium containing a feedstock providing the carbon source, until a recoverable quantity of isobutanol is produced.

Assignees

• Vib Vzw

Inventors

• Dundon Catherine Asleson
• Smith Christopher
• Nahreini Piruz
• Thevelein Johan
• Saerens Sofie

Classifications

• C12N1/16

  Yeasts; Culture media therefor · CPC title

• C12P7/16 Primary

  Butanols · CPC title

• C12N15/81

  for yeasts · CPC title

• C07K14/395

  from Saccharomyces · CPC title

• C12N1/00

  Microorganisms; Compositions thereof (medicinal preparations containing material from protozoa, bacteria or viruses A61K35/66, from algae A61K36/02, from fungi A61K36/06; preparing medicinal bacterial antigen or antibody compositions, e.g. bacterial vaccines, A61K39/00); Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor · CPC title