Methylbutanol as an advanced biofuel
This invention describes genes, metabolic pathways, microbial strains and methods to produce methylbutanol and other compounds of interest from renewable feedstocks.
1. A recombinant microorganism comprising at least one exogenous nucleic acid molecule, wherein the at least one exogenous nucleic acid molecule encodes (i) a pyruvate decarboxylase having at least 90% amino acid sequence identity to a Pichia stipitis pyruvate decarboxylase PDC3-6 gene product of SEQ ID NO:53 having pyruvate decarboxylase activity and (ii) an alcohol dehydrogenase that catalyzes a 2-methylbutanal to 2-methylbutanol conversion; and wherein the recombinant microorganism produces 2-methylbutanol.
2. The recombinant microorganism of claim 1 , wherein the alcohol dehydrogenase is from a genus selected from the group consisting of Saccharomyces, Pichia , or Mycobacterium.
3. The recombinant microorganism of claim 2 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae ADH6 gene product of SEQ ID NO:66 having alcohol dehydrogenase activity.
4. The recombinant microorganism of claim 2 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae SFA1 gene product of SEQ ID NO:76 having alcohol dehydrogenase activity.
5. The recombinant microorganism of claim 1 , wherein the recombinant microorganism is from a genus selected from the group consisting of Escherichia, Pseudomonas, Bacillus, Corynebacterium, Clostridium, Lactobacillus, Pichia , or Saccharomyces.
6. A recombinant microorganism comprising at least one exogenous nucleic acid molecule, wherein the at least one exogenous nucleic acid molecule encodes (i) a pyruvate decarboxylase having at least 90% amino acid sequence identity to a Pichia stipitis pyruvate decarboxylase PDC3-6 gene product of SEQ ID NO:53 having pyruvate decarboxylase activity, wherein said recombinant microorganism produces 2-methylbutanol.
7. The recombinant microorganism of claim 6 , further comprising at least one exogenous nucleic acid molecule encoding an alcohol dehydrogenase that catalyzes a 2-methylbutanal to 2-methylbutanol conversion.
8. The recombinant microorganism of claim 7 , wherein the alcohol dehydrogenase is from a genus selected from the group consisting of Saccharomyces, Pichia , or Mycobacterium.
9. The recombinant microorganism of claim 8 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae ADH6 of SEQ ID NO:66 having alcohol dehydrogenase activity.
10. The recombinant microorganism of claim 8 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae SFA1 of SEQ ID NO:76 having alcohol dehydrogenase activity.
11. A recombinant method for producing a 2-methylbutanol, comprising culturing the recombinant microorganism of claim 1 in a culture medium containing a carbon source, wherein the recombinant microorganism produces spent culture medium from the culture medium by metabolizing the carbon source to the 2-methylbutanol; and recovering the 2-methylbutanol from the spent culture medium.
12. The method of claim 11 , wherein the alcohol dehydrogenase is from a genus selected from the group consisting of Saccharomyces, Pichia , or Mycobacterium.
13. The method of claim 12 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae ADH6 of SEQ ID NO:66 having alcohol dehydrogenase activity.
14. The method of claim 12 , wherein the alcohol dehydrogenase has at least 90% amino acid sequence identity to Saccharomyces cerevisiae SFA1 of SEQ ID NO:76 having alcohol dehydrogenase activity.
15. The method of claim 11 , wherein the recombinant microorganism is selected from a genus selected from the group consisting of Escherichia, Pseudomonas, Bacillus, Corynebacterium, Clostridium, Lactobacillus, Pichia , or Saccharomyces.
16. The method of claim 15 , wherein the recombinant microorganism is Escherichia coli.
17. The method of claim 15 , wherein the recombinant microorganism is Saccharomyces cerevisiae.
18. The method of claim 15 , wherein the recombinant microorganism is Pichia stipitis.
19. A recombinant method for producing a 2-methylbutanol, comprising culturing the recombinant microorganism of claim 6 in a culture medium containing a carbon source, wherein the recombinant microorganism produces spent culture medium from the culture medium by metabolizing the carbon source to the 2-methylbutanol; and recovering the 2-methylbutanol from the spent culture medium.
20. A recombinant method for producing a 2-methylbutanol, comprising culturing the recombinant microorganism of claim 7 in a culture medium containing a carbon source, wherein the recombinant microorganism produces spent culture medium from the culture medium by metabolizing the carbon source to the 2-methylbutanol; and recovering the 2-methylbutanol from the spent culture medium.