Methods of Producing 6-Carbon Monomers From 8-Carbon Compounds
AssigneeInvista North America S.A.R.L.
InventorsAdriana Leonora Botes, Alex Van Eck Conradie, Ramdane Haddouche
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Abstract
This document describes biochemical pathways for producing 6-hydroxyhexanoic acid using a monooxygenase to form a 7-hydroxyoctanoate intermediate, which can be converted to 6-hydroxyhexanoate using a polypeptide having monooxygenase, secondary alcohol dehydrogenase, or esterase activity. 6-hydroxyhexanoic acid can be enzymatically converted to adipic acid, caprolactam, 6-aminohexanoic acid, hexamethylenediamine or 1,6-hexanediol. This document also describes recombinant hosts producing 6-hydroxyhexanoic acid as well as adipic acid, caprolactam, 6-aminohexanoic acid, hexamethylenediamine and 1,6-hexanediol.
Claims43
A method of producing 7-hydroxyoctanoate, said method comprising enzymatically converting octanoate to 7-hydroxyoctanoate using a monooxygenase classified under EC. 1.14.14.1.
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A method for biosynthesizing 6-hydroxyhexanoate, said method comprising either:
- (i) enzymatically synthesizing 7-hydroxyoctanoate from octanoyl-CoA or octanoyl-[acp] using a thioesterase and a monooxygenase classified under EC 1.14.14.1, and enzymatically converting 7-hydroxyoctanoate to 6-hydroxyhexanoate using a secondary alcohol dehydrogenase, a monooxygenase classified under EC 1.14.13.-, and an esterase;
- or
- (ii) enzymatically synthesizing 7-hydroxyoctanoate from 2-oxononanoate using a decarboxylase, an aldehyde dehydrogenase, and a monooxygenase classified under EC 1.14.14.1, and enzymatically converting 7-hydroxyoctanoate to 6-hydroxyhexanoate using a secondary alcohol dehydrogenase, a monooxygenase classified under EC 1.14.13.-, and an esterase;
- wherein said monooxygenase classified under EC 1.14.14.1 has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:
- 18, or is any other monooxygenase classified under EC 1.14.14.1;
- wherein said thioesterase has at least 70% identity to the amino acid sequence set forth in SEQ ID NO:
- 1, 15, 16, or 17, or is any other thioesterase;
- wherein said esterase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:
- 22 or is any other esterase;
- where said monooxygenase classified under EC 1.14.13.- has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:
- 20 or SEQ ID NO:
- 21, or is any other monooxygenase classified under EC 1.14.13;
- wherein said secondary alcohol dehydrogenase has at least 70% identity to the amino acid sequence set forth in SEQ ID NO:
- 19, or is any other secondary alcohol dehydrogenase;
- and/or
- wherein said decarboxylase has at least 70% identity to the amino acid sequence set forth in SEQ ID NO:
- 23, or is any other decarboxylase.
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A recombinant host comprising at least one exogenous nucleic acid encoding (i) a monooxygenase classified under EC 1.14.14.1;
- (ii) a thioesterase, or a decarboxylase and an aldehyde dehydrogenase, (iii) a secondary alcohol dehydrogenase, (iv) a monooxygenase classified under EC 1.14.13.-, and (v) an esterase, said host producing 6-hydroxyhexanoate.
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A non-naturally occurring organism comprising at least one exogenous nucleic acid encoding at least one polypeptide having the activity of at least one enzyme depicted in any one of FIGS. 1 to 5.
A non-naturally occurring biochemical network comprising one or more polypeptides having the activity of a monooxygenase, a secondary alcohol dehydrogenase, and an esterase A.
A nucleic acid construct or expression vector comprising
- (a) a polynucleotide encoding a polypeptide having monooxygenase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having monooxygenase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO:
- 18;
- (b) a polynucleotide encoding a polypeptide having esterase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having esterase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO:
- 22;
- (c) a polynucleotide encoding a polypeptide having thioesterase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having thioesterase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NOs:
- 1, 15, 16, or 17;
- (d) a polynucleotide encoding a polypeptide having decarboxylase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having decarboxylase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO:
- 23;
- (e) a polynucleotide encoding a polypeptide having alcohol dehydrogenase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having alcohol dehydrogenase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO:
- 21;
- (f) a polynucleotide encoding a polypeptide having ω-transaminase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having ω-transaminase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NOs:
- 7-12;
- (g) a polynucleotide encoding a polypeptide having carboxylate reductase activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having carboxylate reductase activity is selected from the group consisting of:
- (a) a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NOs:
- 2-6 or 24;
- or
- (h) a polynucleotide encoding a polypeptide having monooxygenase, primary alcohol dehydrogenase, 6-hydroxyhexanoate dehydrogenase, 7-oxoheptanoate dehydrogenase, 6-oxohexanoate dehydrogenase, 5-oxovalerate dehydrogenase, aldehyde dehydrogenase, 5-hydroxypentanoate dehydrogenase, 4-hydroxybutyrate dehydrogenase, carboxylate reductase, N-acetyltransferase, acetylputrescine deacylase or ω-transaminase activity.
Dependencies are inferred from the claim text.