Genetically modified acetogenic cell

1. An acetogenic microbial cell which is capable of producing at least one higher alcohol from a carbon source, wherein the acetogenic microbial cell is genetically modified to comprise at least one enzyme, E 8 , a butyryl-CoA: acetate CoA transferase (cat3), wherein said E 8 comprises an amino acid sequence having at least 100% sequence identity with the amino acid sequence of SEQ ID NO:1 of an enzyme obtained from Clostridium kluyveri , and wherein the higher alcohol comprises the structure of formula I below and has 4 to 10 carbon atoms:

wherein the acetogenic microbial cell is Clostridium ljungdahlii or Clostridium autothenogenum.

2. The acetogenic microbial cell of claim 1 , wherein the cell is genetically modified to comprise an increased expression relative to its wild type cell of at least one further enzyme selected from the group consisting of E 1 , E 3 to E 7 and E 12 , wherein E 1 is an alcohol dehydrogenase (adh) comprising at least a 90% sequence identity with SEQ ID NO: 18 or SEQ ID NO: 19 and comprises alcohol dehydrogenase activity, E 3 is an acetoacetyl-CoA thiolase (thl) comprising at least a 90% sequence identity with SEQ ID NO: 2 and comprises acetoacetyl-CoA thiolase activity, E 4 is a 3-hydroxybutyryl-CoA dehydrogenase (hbd) comprising at least a 90% sequence identity with SEQ ID NO: 3 and comprises 3-hydroxybutyryl-CoA dehydrogenase activity, E 5 is a 3-hydroxybutyryl-CoA dehydratase (crt) comprising at least a 90% sequence identity with SEQ ID NO: 4 and comprises 3-hydroxybutyryl-CoA dehydratase activity, E 6 is a butyryl-CoA dehydrogenase (bcd) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 5-7 and comprises butyryl-CoA dehydrogenase activity, E 7 is an electron transfer flavoprotein subunit (etf) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 8-13 and comprises electron transfer flavoprotein subunit activity, and E 12 is a trans-2-enoyl-CoA reductase or crotonyl-CoA reductase comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 15-17 and comprises trans-2-enoyl-CoA reductase or crotonyl-CoA reductase activity.

3. The acetogenic microbial cell of claim 1 , wherein the increased expression of the E 8 increases due to an increase in the copy number of a gene for E 8 .

4. The acetogenic microbial cell of claim 2 , wherein:

E 1 comprises at least a 95% sequence identity with SEQ ID NO: 18 or SEQ ID NO: 19 and comprises alcohol dehydrogenase activity;

E 3 comprises at least a 95% sequence identity with SEQ ID NO: 2 and comprises acetoacetyl-CoA thiolase activity;

E 4 comprises at least a 95% sequence identity with SEQ ID NO: 3 and comprises 3-hydroxybutyryl-CoA dehydrogenase activity;

E 5 comprises at least a 95% sequence identity with SEQ ID NO: 4 and comprises 3-hydroxybutyryl-CoA dehydratase activity;

E 6 comprises at least a 95% sequence identity with at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 5-7 and comprises butyryl-CoA dehydrogenase activity;

E 7 comprises at least a 95% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 8-13 and comprises electron transfer flavoprotein subunit activity;

E 12 comprises at least a 95% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NO: 15-17 and comprises trans-2-enoyl-CoA reductase or crotonyl-CoA reductase activity.

5. The acetogenic microbial cell of claim 1 , wherein the higher alcohol is selected from the group consisting of 1-butanol, 2-methyl-1-butanol, isobutanol, 3-methyl-1-butanol, 1-hexanol, 1-octanol, 1-pentanol, 1-heptanol, 3-methyl-1-pentanol, 4-methyl-1-hexanol, 5-methyl-1-heptanol, 4-methyl-1-pentanol, 5-methyl-1-hexanol, 6-methyl-1-heptanol and combinations thereof.

6. A method of producing at least one higher alcohol, the method comprising

contacting the acetogenic microbial cell of claim 1 with a medium comprising a carbon source.

7. The method of claim 6 , wherein the carbon source comprises CO and/or CO 2 .

8. The method of claim 6 , wherein the microbial cell is genetically modified to comprise an increased expression relative to its wild type cell of at least one further enzyme selected from the group consisting of E 1 , E 3 to E 7 and E 12 , wherein E 1 is an alcohol dehydrogenase (adh) comprising at least a 90% sequence identity with SEQ ID NO: 18 or SEQ ID NO: 19, E 3 is an acetoacetyl-CoA thiolase (thl) comprising at least a 90% sequence identity with SEQ ID NO: 2, E 4 is a 3-hydroxybutyryl-CoA dehydrogenase (hbd) comprising at least a 90% sequence identity with SEQ ID NO: 3, E 5 is a 3-hydroxybutyryl-CoA dehydratase (crt) comprising at least a 90% sequence identity with SEQ ID NO: 4, E 6 is a butyryl-CoA dehydrogenase (bcd) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 5-7, E 7 is an electron transfer flavoprotein subunit (etf) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 8-13, and E 12 is a trans-2-enoyl-CoA reductase or crotonyl-CoA reductase comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 15-17.

9. The method of claim 8 , wherein the acetogenic microbial cell is genetically modified to comprise an increased expression relative to its wild type cell of E 3 , E 4 and E 12 .

10. The method of claim 6 , wherein the higher alcohol is selected from the group consisting of 1-butanol, 2-methyl-1-butanol, isobutanol, 3-methyl-1-butanol, 1-hexanol, 1-octanol, 1-pentanol, 1-heptanol, 3-methyl-1-pentanol, 4-methyl-1-hexanol, 5-methyl-1-heptanol, 4-methyl-1-pentanol, 5-methyl-1-hexanol, 6-methyl-1-heptanol and combinations thereof.

11. The method of claim 6 , wherein increased expression of E 8 is due to expression of a heterologous E 8 enzyme, an increase in the copy number of a gene for E 8 , the use of a heterologous promoter, or combinations thereof.

12. An acetogenic microbial cell which is capable of producing at least one higher alcohol from a carbon source, wherein:

the acetogenic microbial cell is genetically modified to comprise increased butyryl-CoA: acetate CoA transferase (cat3 activity) relative to the acetogenic microbial cell before being genetically modified, said increased activity being at least partially due to the enzyme E 8 , wherein said E 8 comprises an amino acid sequence having at least 100% sequence identity with the amino acid sequence of SEQ ID NO: 1 of an enzyme of obtained from Clostridium kluyveri;

the higher alcohol comprises the structure of formula I below and has 4 to 10 carbon atoms:

wherein R=H or CH 3 , and n=1-6;

and wherein the acetogenic microbial cell is Clostridium ljungdahlii or Clostridium autothenogenum.

13. The acetogenic microbial cell of claim 12 , wherein the cell is genetically modified to comprise increased expression relative to its wild type cell of at least one enzyme selected from the group consisting of E 1 , E 3 to E 7 and E 12 , wherein E 1 is an alcohol dehydrogenase (adh) comprising at least a 90% sequence identity with SEQ ID NO: 18 or SEQ ID NO: 19 and comprises alcohol dehydrogenase activity, E 3 is an acetoacetyl-CoA thiolase (thl) comprising at least a 90% sequence identity with SEQ ID NO: 2 and comprises acetoacetyl-CoA thiolase activity, E 4 is a 3-hydroxybutyryl-CoA dehydrogenase (hbd) comprising at least a 90% sequence identity with SEQ ID NO: 3 and comprises 3-hydroxybutyryl-CoA dehydrogenase activity, E 5 is a 3-hydroxybutyryl-CoA dehydratase (crt) comprising at least a 90% sequence identity with SEQ ID NO: 4 and comprises 3-hydroxybutyryl-CoA dehydratase activity, E 6 is a butyryl-CoA dehydrogenase (bcd) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 5-7 and comprises butyryl-CoA dehydrogenase activity, E 7 is an electron transfer flavoprotein subunit (etf) comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 8-13 and comprises electron transfer flavoprotein subunit activity, and E 12 is a trans-2-enoyl-CoA reductase or crotonyl-CoA reductase comprising at least a 90% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 15-17 and comprises trans-2-enoyl-CoA reductase or crotonyl-CoA reductase activity.

14. The acetogenic microbial cell of claim 13 , wherein:

E 1 comprises at least a 95% sequence identity with SEQ ID NO: 18 or SEQ ID NO: 19 and comprises alcohol dehydrogenase activity;

E 3 comprises at least a 95% sequence identity with SEQ ID NO: 2 and comprises acetoacetyl-CoA thiolase activity;

E 4 comprises at least a 95% sequence identity with SEQ ID NO: 3 and comprises 3-hydroxybutyryl-CoA dehydrogenase activity;

E 5 comprises at least a 95% sequence identity with SEQ ID NO: 4 and comprises 3-hydroxybutyryl-CoA dehydratase activity;

E 6 comprises at least a 95% sequence identity with at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 5-7 and comprises butyryl-CoA dehydrogenase activity;

E 7 comprises at least a 95% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 8-13 and comprises electron transfer flavoprotein subunit activity;

E 12 comprises at least a 95% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NO: 15-17 and comprises trans-2-enoyl-CoA reductase or crotonyl-CoA reductase activity.

15. The acetogenic microbial cell of claim 14 , wherein the higher alcohol is selected from the group consisting of 1-butanol, 2-methyl-1-butanol, isobutanol, 3-methyl-1-butanol, 1-hexanol, 1-octanol, 1-pentanol, 1-heptanol, 3-methyl-1-pentanol, 4-methyl-1-hexanol, 5-methyl-1-heptanol, 4-methyl-1-pentanol, 5-methyl-1-hexanol, 6-methyl-1-heptanol and combinations thereof.

16. The acetogenic microbial cell of claim 12 , wherein the higher alcohol is selected from the group consisting of 1-butanol, 2-methyl-1-butanol, isobutanol, 3-methyl-1-butanol, 1-hexanol, 1-octanol, 1-pentanol, 1-heptanol, 3-methyl-1-pentanol, 4-methyl-1-hexanol, 5-methyl-1-heptanol, 4-methyl-1-pentanol, 5-methyl-1-hexanol, 6-methyl-1-heptanol and combinations thereof.