Conversion of S-lignin compounds to useful intermediates
The present disclosure relates to a genetically modified microbial cell that includes a genetic modification resulting in the expression of a vanillate demethylase, where the microbial cell is capable of metabolizing at least one S-lignin decomposition molecule including at least one of syringate and/or 3-O-methyl gallate, and the genetically modified microbial cell is capable of producing gallate. In some embodiments of the present disclosure, the vanillate demethylase may include VanAB.
1. A genetically modified Pseudomonas comprising: a genetic modification resulting in the overexpression of a vanillate demethylase, wherein: the Pseudomonas metabolizes S-lignin decomposition molecules consisting of syringate and 3-O-methyl gallate, and the genetically modified Pseudomonas produces gallate.
2. The genetically modified Pseudomonas of claim 1 , wherein the vanillate demethylase is VanAB.
3. The genetically modified Pseudomonas of claim 1 , wherein the genetically modified Pseudomonas produces at least one of 2-hydroxy-2H-pyran-4,6-dicarboxylic acid (PDC), (1E,3E)-4-hydroxybuta-1,3-diene-1,2,4-tricarboxylic acid, (1E)-4-oxobut-1-ene-1,2,4-tricarboxylic acid, 2-hydroxy-4-oxobutane-1,2,4-tricarboxylic acid, oxaloacetate, or pyruvate.
4. The genetically modified Pseudomonas of claim 1 , further comprising a genetic modification resulting in the expression of a 3,4-dioxygenase.
5. The genetically modified Pseudomonas of claim 4 , wherein the 3,4-dioxygenase is protocatechuate 3,4-dioxygenase (PcaHG).
6. The genetically modified Pseudomonas of claim 5 , further comprising an endogenous genetic deletion that causes a lack of expression of a dioxygenase.
7. The genetically modified Pseudomonas of claim 6 , wherein: the dioxygenase comprises GalA, and the genetically modified Pseudomonas produces PDC.
8. The genetically modified Pseudomonas of claim 1 , wherein the Pseudomonas is P. putida KT2440.
9. A method for making at least one of 2-hydroxy-2H-pyran-4,6-dicarboxylic acid (PDC), (1E,3E)-4-hydroxybuta-1,3-diene-1,2,4-tricarboxylic acid, (1E)-4-oxobut-1-ene-1,2,4-tricarboxylic acid, 2-hydroxy-4-oxobutane-1,2,4-tricarboxylic acid, oxaloacetate, or pyruvate comprising exposing a genetically modified Pseudomonas to a solution containing at least one of S-lignin decomposition molecules, syringate or 3-O-methyl gallate wherein the genetically modified Pseudomonas cell comprises a genetic modification that results in the overexpression of vanillate demethylase.
10. The method of claim 9 wherein the vanillate demethylase is VanAB.
11. The method of claim 9 wherein the genetically modified Pseudomonas further comprises a genetic modification resulting in the expression of a 3,4-dioxygenase.
12. The method of claim 11 , wherein the 3,4-dioxygenase comprises is protocatechuate 3,4-dioxygenase (PcaHG).
13. The method of claim 9 wherein the genetically modified Pseudomonas further comprises an endogenous genetic deletion that causes a lack of expression of a dioxygenase.
14. The method of claim 13 wherein the dioxygenase comprises GalA, and the genetically modified Pseudomonas produces PDC.
15. The method of claim 14 wherein the modified Pseudomonas produces PDC at a concentration of up to 3.38 mM.
16. The method of claim 15 wherein the modified Pseudomonas produces up to 3.38 mM PDC after about 72 hours of growth at a yield of up to 68%.