Acyl-ACP thioesterase genes and uses therefor
The present invention provides novel genes encoding Class II acyl-ACP thioesterases and variants thereof that are active on C8, C10, C12, C14, C16, and C18 acyl-ACP substrates. The thioesterases can be introduced into transgenic organisms, including microorganisms and photosynthetic organisms, for producing fatty acids and fatty acid products.
1. A transgenic organism comprising an exogenous nucleic acid molecule encoding a Class II acyl-ACP thioesterase comprising an amino acid sequence selected from the group consisting of an amino acid sequence having at least 90% identity to amino acid 64 to amino acid 361 of SEQ ID NO:51; the amino acid sequence of amino acid 66 to amino acid 362 of SEQ ID NO:55; an amino acid sequence having at least 96% identity to amino acid 65 to amino acid 356 of SEQ ID NO:69; and the amino acid sequence of amino acid 65 to amino acid 362 of SEQ ID NO:85.
2. A transgenic organism according to claim 1 , wherein the acyl-ACP thioesterase has a C12, C14, and/or C16 acyl substrate preference.
3. A transgenic organism according to claim 1 , wherein the transgenic organism further comprises at least one exogenous nucleic acid molecule encoding an acetyl-CoA carboxylase, a ketoacyl-CoA synthase, a fatty acid elongase, an acyl-CoA synthetase, a fatty acyl-CoA reductase, a fatty aldehyde reductase, an alcohol acetyl transferase, an acyl-CoA alcohol transacylase, an acyltransferase, a wax synthase, an aldehyde decarbonylase, or a fatty acid decarboxylase.
4. A transgenic organism according to claim 1 , wherein the organism is plant.
5. A transgenic organism according to claim 1 , wherein the organism is a microorganism.
6. A transgenic organism according to claim 5 , wherein the organism is a photosynthetic microorganism.
7. A transgenic organism according to claim 6 , wherein the photosynthetic microorganism is a eukaryotic alga.
8. A transgenic organism according to claim 6 , wherein the photosynthetic microorganism is a cyanobacterium.
9. A method of making a fatty acid product, comprising:
culturing a transgenic microorganism according to claim 5 ; and
isolating a fatty acid product from the organism or culture medium.
10. The method of claim 9 , wherein the fatty acid product is a free fatty acid.
11. The method of claim 9 , wherein the fatty acid product comprises one or more triglycerides, fatty aldehydes, fatty alcohols, fatty esters, hydrocarbons, or fatty acids.
12. The method of claim 9 , wherein the microorganism is a photosynthetic microorganism.
13. The method of claim 12 , wherein the microorganism is cultured mixotrophically.
14. The method of claim 12 , wherein the microorganism is cultured phototrophically.
15. The method of claim 12 , wherein the photosynthetic organism is a eukaryotic alga.
16. The method of claim 12 , wherein the photosynthetic organism is a cyanobacterium.
17. The method of claim 9 , wherein at least 50% of the fatty products isolated from the organism, the culture medium, or both are C12 fatty acid products, C14 fatty acid products, C16 fatty acid products, or a combination thereof.
18. The method of claim 9 , wherein at least 20% of the fatty products isolated from the organism, the culture medium, or both are C14 fatty acid products.
19. A vector comprising a recombinant nucleic acid molecule encoding a Class II acyl-ACP thioesterase comprising an amino acid sequence selected from the group consisting of:
a) an amino acid sequence having at least 90% identity to amino acid 64 to amino acid 361 of SEQ ID NO:51;
b) the amino acid sequence of amino acid 66 to amino acid 362 of SEQ ID NO:55;
c) an amino acid sequence having at least 96% identity to amino acid 65 to amino acid 356 of SEQ ID NO:69; and
d) the amino acid sequence of amino acid 65 to amino acid 362 of SEQ ID NO:85;
wherein the vector further comprises a selectable marker.
20. A vector according to claim 19 , encoding a Class II acyl-ACP thioesterase comprising a nucleic acid sequence encoding a thioesterase that comprises an amino acid sequence selected from the group consisting of:
an amino acid sequence having at least 90% identity to SEQ ID NO:51;
the amino acid sequence of SEQ ID NO:55;
an amino acid sequence having at least 96% identity to SEQ ID NO:69; and
the amino acid sequence of SEQ ID NO:85.