Biological synthesis of difunctional alkanes from carbohydrate feedstocks

Abstract

Aspects of the invention relate to methods for the production of difunctional alkanes in host cells. In particular, aspects of the invention describe components of genes associated with the difunctional alkane production from carbohydrate feedstocks in host cells. More specifically, aspects of the invention describe metabolic pathways for the production of adipic acid, aminocaproic acid, caprolactam, and hexamethylenediamine via 2-ketopimelic acid.

What is claimed is: 
     
         1 . A method of producing a α,ω difunctional Cn alkane from α-ketoglutarate wherein α and ω terminal functional groups are selected from the group of —OH, —COOH and —NH3 and wherein n is an integer in the range of 4 to 7 comprising:
 culturing a metabolically engineered host cell under conditions sufficient to produce the α,ω difunctional Cn alkane from a α-ketoacid, wherein the metabolically engineered host cell is genetically modified with a nucleic acid comprising at least one nucleotide sequence encoding at least two biosynthetic pathway enzymes; and 
 separating said α,ω difunctional Cn alkane. 
 
     
     
         2 . The method of  claim 1  wherein in the step of culturing, the engineered host cell produces α-ketoadipate, α-ketopimelate, α-ketosuberate or a combination thereof from α-ketoglutarate. 
     
     
         3 . The method of  claim 1  wherein the nucleic acid comprises one or more nucleic acid sequences encoding a set of at least two enzymes of a biosynthetic pathway, comprising a recombinant α-ketoacid decarboxylase capable of using the α-ketoacid as a substrate and at least a recombinant aldehyde dehydrogenase, a recombinant alcohol dehydrogenase, a recombinant aminotransferase, or a combination thereof. 
     
     
         4 . The method of  claim 1  wherein the nucleic acid comprises one or more nucleic acid sequences encoding a set of at least two enzymes of a biosynthetic pathway, comprising a recombinant 2-aminotransferase capable of using the α-ketoacid as a substrate and at least a recombinant aminodecarboxylase, a recombinant reductase, a recombinant dehydrogenase, a recombinant decarboxylase, a recombinant aldehyde dehydrogenase, a recombinant alcohol dehydrogenase, a recombinant aminotransferase, a recombinant aminoaldehyde dehydrogenase, a recombinant 2-aminoaldehyde mutase, a recombinant dehydratase or a combination thereof. 
     
     
         5 . The method of  claim 1  wherein the nucleic acid comprises one or more nucleic acid sequences encoding a set of at least two enzymes of a biosynthetic pathway, comprising a recombinant dehydrogenase capable of using the α-ketoacid as a substrate and at least a recombinant dehydratase, a recombinant aldehyde dehydrogenase, a recombinant alcohol dehydrogenase, a recombinant 1-aminotransferase, or a combination thereof. 
     
     
         6 . The method of  claim 1  wherein the at least one nucleotide acid sequence encodes at least one polypeptide selected from a α-ketoacid decarboxylase (EC 4.1.1.-), a dehydrogenase (EC 1.1.1.-) and combination thereof. 
     
     
         7 . The method of  claim 1  wherein the at least one nucleotide acid sequence encodes at least one polypeptide selected from a α-ketoacid decarboxylase (EC 4.1.1.-), an amino transferase (EC 1.4.1.-), an amino transferase (EC 2.6.1.-) and combination thereof. 
     
     
         8 . The method of  claim 7  further comprising at least one nucleotide acid sequence further encodes a recombinant aldehyde dehydrogenase (EC 1.2.1.3). 
     
     
         9 . The method of  claim 1  wherein the at least one nucleotide acid sequence encodes at least one polypeptide selected from a α-aminoadipate-aminotransferase (EC 2.6.1.39), a diaminopimelate dehydrogenase (EC 1.4.1.16), an amino adipate reductase (EC 1.2.1.31), a saccharopine dehydrogenase (EC 1.5.1.-), a lysine decarboxylase (EC 4.1.1.18), an ornithine decarboxylase (EC4.1.1.17) and combination thereof. 
     
     
         10 . The method of  claim 1  wherein the α,ω difunctional Cn alkane is a Cn dicarboxylic acid, Cn hydroxycarboxylic acid or Cn alkane diol. 
     
     
         11 . The method of  claim 1  wherein the α,ω difunctional Cn alkane is one of 1,4-butane diol, 1-hydroxybutanoate, succinic acid, 1,4-diaminobutane, 4-aminobutanal and 4-aminobutanol. 
     
     
         12 . The method of  claim 1  wherein the α,ω difunctional Cn alkane is one of 1-hydroxypentanoate, 1,5-pentanediol, glutarate, pentane-1,5-diamine, 5-aminopentanal and 5-aminopentanol. 
     
     
         13 . The method of  claim 1  wherein the α,ω difunctional Cn alkane is one of 1-hydroxyhexanoate, 1,6-hexanediol, adipate, hexamethylenediamine, amino caproic acid, 6-amino hexanal and 6-aminohexanol. 
     
     
         14 . The method of  claim 1  wherein the α,ω difunctional Cn alkane is one of 1-hydroxyheptanoate, 1,7-heptanediol, pimelic acid, 1,7-diaminoheptane, 7-aminoheptanal and 7-aminoheptanol. 
     
     
         15 . The method of  claim 1  wherein, in the step of culturing, the engineered host cell further comprises one or more nucleic acid sequences encoding a recombinant homocitrate synthase, a recombinant homoaconitase and a recombinant homoisocitrate dehydrogenase, or homologs thereof. 
     
     
         16 . The method of  claim 1  wherein the at least one nucleotide acid sequence encodes at least one polypeptide selected from an aminotransferase (EC 2.6.1.7), an aldehyde dehydrogenase (EC 1.2.1.3), a glutamate semialdehyde mutase (EC 5.4.3.8), an 3-oxoacyl-[acyl-carrier-protein]reductase (EC 1.1.1.100), a fatty acid synthase (EC 2.3.1.-), a dehydratase (EC 4.2.1.59), a 3-hydroxyoctanoyl-[acyl-carrier-protein]dehydratase (EC 4.2.1.59), an enoyl-[acyl-carrier-protein]reductase (EC 1.3.1.9) and combination thereof.