Producing biofuels using polyketide synthases
The present invention provides for a non-naturally occurring polyketide synthase (PKS) capable of synthesizing a carboxylic acid or a lactone, and a composition such that a carboxylic acid or lactone is included. The carboxylic acid or lactone, or derivative thereof, is useful as a biofuel. The present invention also provides for a recombinant nucleic acid or vector that encodes such a PKS, and host cells which also have such a recombinant nucleic acid or vector. The present invention also provides for a method of producing such carboxylic acids or lactones using such a PKS.
1. A method of producing a carboxylic acid of Formula (I),
wherein R 1 is H or —CH 3 ; R 2 is H, —CH 3 , or —CH 2 CH 3 ; and R 3 is H, —CH 3 , or —CH 2 CH 3 ; or R 2 and R 3 are each H, and R 1 is H, —CH 3 , or —(CH 2 ) n CH 3 , where n is an integer from 1 to 6; and
wherein the method comprises culturing a host cell comprising a vector encoding a non-naturally occurring polyketide synthase in a culture medium under conditions where the PKS is expressed and the carboxylic acid is produced, wherein the PKS comprises a loading domain and two or more extender modules, and the PKS is a hybrid PKS comprising a combination of modules which in nature are not found in this combination.
2. The method of claim 1 , wherein the carboxylic acid has a structure where R 1 is H or —CH 3 ; R 2 is H, —CH 3 , or —CH 2 CH 3 ; and R 3 is H, —CH 3 , or —CH 2 CH 3 .
3. The method of claim 1 , wherein the carboxylic acid has a structure where R 2 and R 3 are each H, and R 1 is H, —CH 3 , or —(CH 2 ) n CH 3 , where n is an integer from 1 to 6.
4. The method of claim 3 , wherein the carboxylic acid has a structure where R 2 and R 3 are each H, and R 1 is —(CH 2 ) n CH 3 , where n is an integer from 1 to 6.
5. The method of claim 1 , wherein the carboxylic acid has a structure disclosed in Tables 1 and 2.
6. The method of claim 1 , further comprising: (c) isolating said carboxylic acid from the host cell and/or the culture medium.
7. The method of claim 6 , further comprising: (d) reacting the isolated carboxylic acid with an alcohol to produce an ester.
8. A method of producing a lactone of Formula (II),
wherein R 4 is H or —CH 3 ; and R 5 and R 6 are each independently H, —CH 3 , or —CH 2 CH 3 ; and m is 1, 3, 5, 7, or 9; with the proviso that when m is 3, 5, 7, or 9, then R 4 is H or —CH 3 and R 5 and R 6 are each H; and
wherein the method comprises culturing a host cell comprising a vector encoding a non-naturally occurring polyketide synthase in a culture medium under conditions where the PKS is expressed and the lactone is produced, wherein the PKS comprises a loading domain, two or more extender modules, and a thioesterase domain positioned 3′ to the terminal extender module, and the PKS is a hybrid PKS comprising a combination of modules which in nature are not found in this combination.
9. The method of claim 8 , wherein the lactone has a structure where m is 1 and R 5 and R 6 are each independently H, —CH 3 , or —CH 2 CH 3 .
10. The method of claim 8 , wherein the lactone has a structure where m is 3, 5, 7, or 9; R 4 is H or —CH 3 , and R 5 and R 6 are each H.
11. The method of claim 8 , wherein the lactone is any lactone disclosed in Tables 3-7.
12. The method of claim 8 , wherein the lactone is a six-membered lactone having the following chemical structure:
wherein R 1 is H or —CH 3 ; and R 2 and R 3 are each independently H, —CH 3 , or —CH 2 CH 3 .
13. The method of claim 8 , wherein the lactone is an eight-membered lactone having the following chemical structure:
wherein R is H or —CH 3 .
14. The method of claim 8 , wherein the lactone is a ten-membered lactone having the following chemical structure:
wherein R is H or —CH 3 .
15. The method of claim 8 , wherein the lactone is a twelve-membered lactone having the following chemical structure:
wherein R is H or —CH 3 .
16. The method of claim 8 , wherein the lactone is a four-teenmember lactone having the following chemical structure:
wherein R is H or —CH 3 .
17. The method of claim 8 , further comprising: (c) isolating said lactone from the host cell and/or the culture medium.