Microorganisms and methods for the production of butadiene using acetyl-coA

1. A non-naturally occurring microbial organism, said microbial organism having a butadiene pathway and comprising at least four exogenous nucleic acids encoding butadiene pathway enzyme expressed in a sufficient amount to produce butadiene, wherein said butadiene pathway comprises a pathway selected from:

(9) 1B, 1C, 1G, 1I, 1L, 1M, and 1F;

(10) 1B, 1C, 1G, 1I, 1L, 1N, and 1F;

(11) 1B, 1C, 1H, 1I, 1L, 1M, and 1F;

(12) 1B, 1C, 1H, 1I, 1L, 1N, and 1F;

(13) 1B, 1C, 1D, 1J, 1L, 1M, and 1F;

(14) 1B, 1C, 1D, 1J, 1L, 1N, and 1F;

(15) 1B, 1C, 1D, 1K, 1L, 1M, and 1F; and

(16) 1B, 1C, 1D, 1K, 1L, 1N, and 1F,

wherein 1B is a 4-hydroxy 2-oxovalerate aldolase, wherein 1C is a 4-hydroxy 2-oxovalerate dehydratase, wherein 1D is a 2-oxopentenoate reductase, wherein 1F is a 2,4-pentadienoate decarboxylase, wherein 1G is a 2-oxopentenoate ligase, wherein 1H is a 2-oxopentenoate:acetyl CoA CoA transferase, wherein 1I is a 2-oxopentenoyl-CoA reductase, wherein 1J is a 2-hydroxypentenoate ligase, wherein 1K is a 2-hydroxypentenoate:acetyl-CoA CoA transferase, wherein 1L is a 2-hydroxypentenoyl-CoA dehydratase, wherein 1M is a 2,4-Pentadienoyl-CoA hydrolase, and wherein 1N is a 2,4-Pentadienoyl-CoA:acetyl CoA CoA transferase.

2. The non-naturally occurring microbial organism of claim 1 , wherein said microbial organism comprises one, two, three, four, five, six, seven, eight, nine, ten, or eleven exogenous nucleic acids each encoding a butadiene pathway enzyme, or wherein said microbial organism comprises exogenous nucleic acids encoding each of the enzymes of at least one of the pathways selected from (9)-(16).

3. The non-naturally occurring microbial organism of claim 1 further comprising an acetyl-CoA pathway, wherein said acetyl-CoA pathway comprises a pathway selected from:

(1) 3T and 3V;

(2) 3T, 3W, and 3X;

(3) 3U and 3V;

(4) 3U, 3W, and 3X

wherein 3T is a fructose-6-phosphate phosphoketolase, wherein 3U is a xylulose-5-phosphate phosphoketolase, wherein 3V is a phosphotransacetylase, wherein 3W is an acetate kinase, wherein 3X is an acetyl-CoA transferase, an acetyl-CoA synthetase, or an acetyl-CoA ligase.

4. The non-naturally occurring microbial organism of claim 1 , wherein said microbial organism further comprises a formaldehyde fixation pathway, wherein said formaldehyde fixation pathway comprises:

(1) 3D and 3Z;

(2) 3D; or

(3) 3B and 3C,

wherein 3B is a 3-hexulose-6-phosphate synthase, wherein 3C is a 6-phospho-3-hexuloisomerase, wherein 3D is a dihydroxyacetone synthase, wherein 3Z is a fructose-6-phosphate aldolase.

5. The non-naturally occurring microbial organism of claim 1 , wherein said microbial organism further comprises a methanol metabolic pathway, wherein said methanol metabolic pathway comprises a pathway selected from:

(1) 4A and 4B;

(2) 4A, 4B and 4C;

(3) 4J;

(4) 4J, 4K and 4C;

(5) 4J, 4M, and 4N;

(6) 4J and 4L;

(7) 4J, 4L, and 4G;

(8) 4J, 4L, and 4I;

(9) 4A, 4B, 4C, 4D, and 4E;

(10) 4A, 4B, 4C, 4D, and 4F;

(11) 4J, 4K, 4C, 4D, and 4E;

(12) 4J, 4K, 4C, 4D, and 4F;

(13) 4J, 4M, 4N, and 4O;

(14) 4A, 4B, 4C, 4D, 4E, and 4G;

(15) 4A, 4B, 4C, 4D, 4F, and 4G;

(16) 4J, 4K, 4C, 4D, 4E, and 4G;

(17) 4J, 4K, 4C, 4D, 4F, and 4G;

(18) 4J, 4M, 4N, 4O, and 4G;

(19) 4A, 4B, 4C, 4D, 4E, and 4I;

(20) 4A, 4B, 4C, 4D, 4F, and 4I;

(21) 4J, 4K, 4C, 4D, 4E, and 4I;

(22) 4J, 4K, 4C, 4D, 4F, and 4I; and

(23) 4J, 4M, 4N, 4O, and 4I,

wherein 4A is a methanol methyltransferase, wherein 4B is a methylenetetrahydrofolate reductase, wherein 4C is a methylenetetrahydrofolate dehydrogenase, wherein 4D is a methenyltetrahydrofolate cyclohydrolase, wherein 4E is a formyltetrahydrofolate deformylase, wherein 4F is a formyltetrahydrofolate synthetase, wherein 4G is a formate hydrogen lyase, wherein 4I is a formate dehydrogenase, wherein 4J is a methanol dehydrogenase, wherein 4K is a formaldehyde activating enzyme or spontaneous, wherein 4L is a formaldehyde dehydrogenase, wherein 4M is a S-(hydroxymethyl)glutathione synthase or spontaneous, wherein 4N is a glutathione-dependent formaldehyde dehydrogenase, wherein 4O is a S-formylglutathione hydrolase.

6. The non-naturally occurring microbial organism of claim 1 , wherein said microbial organism further comprises a formate assimilation pathway, wherein said formate assimilation pathway comprises a pathway selected from:

(1) 3E;

(2) 3F, and 3G;

(3) 3H, 3I, 3J, and 3K;

(4) 3H, 3I, 3J, 3L, 3M, and 3N;

(5) 3E, 3H, 3I, 3J, 3L, 3M, and 3N;

(6) 3F, 3G, 3H, 3I, 3J, 3L, 3M, and 3N;

(7) 3K, 3H, 3I, 3J, 3L, 3M, and 3N; and

(8) 3H, 3I, 3J, 3O, and 3P,

wherein 3E is a formate reductase, 3F is a formate ligase, a formate transferase, or a formate synthetase, wherein 3G is a formyl-CoA reductase, wherein 3H is a formyltetrahydrofolate synthetase, wherein 3I is a methenyltetrahydrofolate cyclohydrolase, wherein 3J is a methylenetetrahydrofolate dehydrogenase, wherein 3K is a formaldehyde-forming enzyme or spontaneous, wherein 3L is a glycine cleavage system, wherein 3M is a serine hydroxymethyltransferase, wherein 3N is a serine deaminase, wherein 3O is a methylenetetrahydrofolate reductase, wherein 3P is an acetyl-CoA synthase.

7. The non-naturally occurring microbial organism of claim 6 , wherein said formate assimilation pathway further comprises:

(1) 3Q;

(2) 3R and 3S;

(3) 3Y and 3Q; or

(4) 3Y, 3R, and 3S,

wherein 3Q is a pyruvate formate lyase, wherein 3R is a pyruvate dehydrogenase, a pyruvate ferredoxin oxidoreductase, or a pyruvate:NADP+oxidoreductase, wherein 3S is a formate dehydrogenase, wherein 3Y is a glyceraldehyde-3-phosphate dehydrogenase or an enzyme of lower glycolysis.

8. The non-naturally occurring microbial organism of claim 1 , wherein said organism further comprises:

(a) a methanol oxidation pathway, wherein said methanol oxidation pathway comprises 3A, wherein 3A a methanol dehydrogenase;

(b) a carbon monoxide dehydrogenase;

(c) a hydrogenase;

(d) attenuation of one or more endogenous enzymes selected from DHA kinase, methanol oxidase, PQQ-dependent methanol dehydrogenase, DHA synthase or any combination thereof;

(e) attenuation of one or more endogenous enzymes of a competing formaldehyde assimilation or dissimilation pathway;

(f) a gene disruption of one or more endogenous nucleic acids encoding enzymes selected from DHA kinase, methanol oxidase, PQQ-dependent methanol dehydrogenase, DHA synthase or any combination thereof,

(g) a gene disruption of one or more endogenous nucleic acids encoding enzymes of a competing formaldehyde assimilation or dissimilation pathway; or

(h) a hydrogen synthesis pathway catalyzing the synthesis of hydrogen from a reducing equivalent, said hydrogen synthesis pathway comprising an enzyme selected from the group consisting of a hydrogenase, a formate-hydrogene lyase and ferredoxin: NADP+ oxidoreductase.

9. The non-naturally occurring microbial organism of claim 1 , wherein said at least one exogenous nucleic acid is a heterologous nucleic acid.

10. The non-naturally occurring microbial organism of claim 1 , wherein said non-naturally occurring microbial organism is in a substantially anaerobic culture medium.

11. The non-naturally occurring microbial organism of claim 1 , wherein said microbial organism is a species of bacteria, yeast, or fungus.

12. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1G, 1I, 1L, 1M, and 1F.

13. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1G, 1I, 1L, 1N, and 1F.

14. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1H, 1I, 1L, 1M, and 1F.

15. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1H, 1I, 1L, 1N, and 1F.

16. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1D, 1J, 1L, 1M, and 1F.

17. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1D, 1J, 1L, 1N, and 1F.

18. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1D, 1K, 1L, 1M, and 1F.

19. The non-naturally occurring microbial organism of claim 1 , wherein said butadiene pathway comprises 1B, 1C, 1D, 1K, 1L, 1N, and 1F.

20. A method for producing (a) butadiene or (b) butadiene and hydrogen, comprising culturing the non-naturally occurring microbial organism of claim 1 under conditions and for a sufficient period of time to produce butadiene.

21. The method of claim 20 , wherein said method further comprises separating the butadiene or the butadiene and hydrogen from other components in the culture.

22. The method of claim 21 , wherein the separating comprises extraction, continuous liquid-liquid extraction, pervaporation, membrane filtration, membrane separation, reverse osmosis, electrodialysis, distillation, crystallization, centrifugation, extractive filtration, ion exchange chromatography, absorption chromatography, or ultrafiltration.

23. A culture medium comprising bioderived butadiene, wherein said culture medium is separated from a non-naturally occurring microbial organism having the butadiene pathway in claim 1 .