Microorganisms and methods for producing 2-pyrone-4,6-dicarboxylic acid and other compounds
Recombinant microorganisms configured for enhanced production of compounds such as 2-pyrone-4,6-dicarboxylic acid (PDC) and methods of using the recombinant microorganisms for the production of these compounds. The recombinant microorganisms include one or more modifications that reduce 2-pyrone-4,6-dicarboxylic acid (PDC) hydrolase activity, 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) cis-trans isomerase activity, 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) methyl esterase activity, and/or vanillate/3-O-methylgallate O-demethylase activity. The recombinant microorganisms can be used to generate PDC from media comprising plant-derived phenolics, such as syringyl phenolics, guaiacyl phenolics, and p-hydroxyphenyl phenolics. The plant-derived phenolics can be derived from pretreated lignin, including depolymerized lignin or other chemically altered lignin.
1. A recombinant microorganism comprising modifications with respect to a corresponding native microorganism not comprising the modifications, wherein:
the native microorganism comprises a native 2-pyrone-4,6-dicarboxylic acid (PDC) hydrolase gene, a native 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) cis-trans isomerase gene, and a native 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) methyl esterase gene;
the modifications comprise:
a mutation of the native 2-pyrone-4,6-dicarboxylic acid (PDC) hydrolase gene that reduces PDC hydrolase activity in the recombinant microorganism with respect to the corresponding native microorganism;
a mutation of the native 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) cis-trans isomerase gene that reduces CHMOD cis-trans isomerase activity in the recombinant microorganism with respect to the corresponding native microorganism; and
a mutation of the native 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) methyl esterase gene that reduces CHMOD methyl esterase activity in the recombinant microorganism with respect to the corresponding native microorganism; and
the recombinant microorganism is capable of producing 2-pyrone-4,6-dicarboxylic acid.
2. The recombinant microorganism of claim 1 , wherein the modifications further comprise a mutation of a native vanillate/3-O-methylgallate O-demethylase gene present in the corresponding native microorganism, wherein the mutation of the native vanillate/3-O-methylgallate O-demethylase gene reduces vanillate/3-O-methylgallate O-demethylase activity in the recombinant microorganism with respect to the corresponding native microorganism.
3. The recombinant microorganism of claim 1 , wherein the recombinant microorganism is a prokaryote.
4. The recombinant microorganism of claim 1 , wherein the recombinant microorganism is a bacterium.
5. The recombinant microorganism of claim 1 , wherein the recombinant microorganism is a member of Sphingomonadales or Pseudomonadales.
6. The recombinant microorganism of claim 1 , wherein the recombinant microorganism is from the genus Novosphingobium.
7. The recombinant microorganism of claim 1 , wherein the recombinant microorganism exhibits enhanced production of 2-pyrone-4,6-dicarboxylic acid with respect to the corresponding native microorganism.
8. A method for producing 2-pyrone-4,6-dicarboxylic acid comprising culturing the recombinant microorganism as recited in claim 1 in a medium comprising a plant-derived phenolic.
9. The method of claim 8 , wherein the plant-derived phenolic comprises a phenolic selected from the group consisting of a syringyl phenolic, a guaiacyl phenolic, and ap-hydroxyphenyl phenolic.
10. The method of claim 8 , wherein the medium comprises depolymerized lignin comprising the plant-derived phenolic.
11. The method of claim 8 , wherein the medium comprises chemically depolymerized lignin comprising the plant-derived phenolic.
12. The method of claim 8 , further comprising isolating the 2-pyrone-4,6-dicarboxylic acid from the medium and/or the recombinant microorganism.