Production of isopropanol by improved recombinant strains

Patent metadata

Field	Value
Publication number	US-9399781-B2
Application number	US-201214350923-A
Country	US
Kind code	B2
Filing date	Oct 3, 2012
Priority date	Oct 11, 2011
Publication date	Jul 26, 2016
Grant date	Jul 26, 2016

Abstract

The expression vector includes: the nucleic acids coding for the polypeptides forming a polypeptide complex having an enzyme activity allowing acetoacetyl-CoA to be converted to acetoacetate; optionally, at least one nucleic acid coding for a polypeptide having an enzyme activity allowing acetoacetate to be converted to acetone; and at least one nucleic acid coding for a polypeptide having an enzyme activity allowing acetone to be converted to isopropanol; the expression of the nucleic acids being controlled by a single constitutive promoter located upstream of the abovementioned nucleic acids.

First claim

The invention claimed is:

1. A vector comprising:
a nucleic acid represented by the sequence SEQ ID NO: 1, or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 1, coding for subunit A of the acetoacetyl-CoA transferase of C. acetobutylicum and
a nucleic acid represented by the sequence SEQ ID NO: 2 or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 2, coding for subunit B of the acetoacetyl-CoA transferase of C. acetobutylicum , and
optionally a nucleic acid represented by the sequence SEQ ID NO: 4 or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 4, coding for the acetoacetate decarboxylase of C. acetobutylicum , and
a nucleic acid represented by the sequence SEQ ID NO: 3 or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 3, coding for the secondary alcohol dehydrogenase of C. beijerinckii,
the expression of said nucleic acids being controlled by a single constitutive promoter located upstream of the abovementioned nucleic acids, said promoter being selected from the promoter represented by the sequence SEQ ID NO: 5(thl promoter) and the promoter represented by the sequence SEQ ID NO: 6.

2. The expression vector according to claim 1 , wherein said vector comprises:
a nucleic acid represented by the sequence SEQ ID NO: 1,
a nucleic acid represented by the sequence SEQ ID NO: 2,
a nucleic acid represented by the sequence SEQ ID NO: 3,
a nucleic acid represented by the sequence SEQ ID NO: 4, and
a promoter represented by the sequence SEQ ID NO: 5.

3. The expression vector according to claim 1 , wherein said vector comprises:
a nucleic acid represented by the sequence SEQ ID NO: 1,
a nucleic acid represented by the sequence SEQ ID NO: 2,
a nucleic acid represented by the sequence SEQ ID NO: 3, and
a promoter represented by the sequence SEQ ID NO: 5.

4. A microorganism comprising the expression vector according to claim 1 .

5. The microorganism according to claim 4 , wherein said microorganism is a bacteria of the genera selected from the group consisting of Clostridium, Bacillus , and enterobacteria.

6. The microorganism according to claim 5 , wherein said microorganism is selected from the group consisting of Clostridium acetobutylicum, Clostridium beijerinckii, Clostridium saccharoperbutylacetonicum , and Clostridium saccharobutylicum.

7. The microorganism according to claim 5 , wherein said microorganism is E. coli.

8. A method for the production of isopropanol, and/or of D and L 2,3-butanediol, said method comprising:
cultivating a microorganism according to claim 4 , in a culture medium comprising at least one carbon source and one nitrogen source, under conditions allowing the production of isopropanol, and
recovering the isopropanol, and/or D and L 2,3-butanediol, from the culture medium.

9. The expression vector according to claim 1 , wherein the expression vector does comprise the nucleic acid represented by the sequence SEQ ID NO: 4 or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 4, coding for the acetoacetate decarboxylase of C. acetobutylicum.

10. The expression vector according to claim 1 , wherein the expression vector does not comprise the nucleic acid represented by the sequence SEQ ID NO: 4 or a nucleic acid having a sequence identity of at least 85% with the sequence SEQ ID NO: 4, coding for the acetoacetate decarboxylase of C. acetobutylicum.

Assignees

• Ifp Energies Now
• Stichting Dienst Landbouwkundig Onderzoek

Inventors

• Collas Florent
• Marchal Remy
• Clement Benjamin
• Lopez Contreras Ana Maria
• Claassen Pieternel A M

Classifications

• C12P7/52

  Propionic acid; Butyric acids · CPC title

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title

• Y02P20/52

  using catalysts, e.g. selective catalysts · CPC title

• C12N9/88

  Lyases (4.) · CPC title

• Y02E50/10

  Biofuels, e.g. bio-diesel · CPC title