US 2015/0361458 A1 · METHODS, REAGENTS AND CELLS FOR BIOSYNTHESIZING COMPOUND

Abstract

This document describes biochemical pathways for producing 2(E)-heptenedioyl-CoA methyl ester from precursors such as 2-oxo-glutarate, acetyl-CoA, or succinyl-CoA using one or more of a fatty acid O-methyltransferase, a thioesterase, a CoA-transferase, a CoA ligase, as well as recombinant hosts expressing one or more of such enzymes. 2(E)-heptenedioyl-CoA methyl ester can be enzymatically converted to pimeloyl-CoA using a trans-2-enoyl-CoA reductase, and a methylesterase. Pimeloyl-CoA can be enzymatically converted to pimelic acid, 7-aminoheptanoate, 7-hydroxyheptanoate, heptamethylenediamine, or 1,7-heptanediol.

Claims65

Dependent claims

A method of shielding a carbon chain aliphatic backbone, functionalized with terminal carboxyl groups, in a recombinant host, said method comprising:

enzymatically converting a n-carboxy-2-enoic acid to a n-carboxy-2-enoate methyl ester in said host using a polypeptide having the activity of a fatty acid O-methyltransferase, wherein n+1 reflects length of the carbon chain aliphatic backbone;
wherein when the n-carboxy-2-enoic acid is 2(E) heptenedioic acid, the method optionally further comprises enzymatically converting 2(E)-heptenedioate methyl ester to pimeloyl-CoA.

A recombinant host comprising an exogenous nucleic acid encoding a polypeptide having fatty acid O-methyltransferase activity, said host producing 2(E)-heptenedioate methyl ester;

wherein optionally:
the host further comprises an exogenous polypeptide having thioesterase or CoA-transferase activity and the host produces 2(E)-heptenedioate;
the host further comprises an exogenous polypeptide having CoA ligase activity and the host produces 2(E)-heptendioyl-CoA methyl ester;
host further comprises an exogenous polypeptide having trans-2-enoyl-CoA reductase activity and the host produces pimeloyl-CoA, adipyl-CoA methyl ester and/or an exogenous polypeptide having pimeloyl-[acp]methyl ester methylesterase activity;
the host further comprises one or more exogenous polypeptides having homocitrate synthase, homocitrate dehydratase, homoaconitate hydratase, isohomocitrate dehydrogenase, decarboxylase, indolepyruvate decarboxylase, glutarate-semialdehyde dehydrogenase, or a glutarate:CoA ligase activity;
the host further comprises one or more exogenous polypeptides selected from the group consisting of polypeptides having (a) β-ketothiolase activity or acetyl-carboxylase activity in combination with acetoacetyl-CoA synthase activity, (b) 3-hydroxybutyryl-CoA dehydrogenase activity, (c) enoyl-CoA hydratase activity, and either (d) glutaryl-CoA dehydrogenase activity in combination with enoyl-CoA reductase activity or trans-2-enoyl-CoA reductase activity or (e) glutaconyl-CoA decarboxylase activity;
the host further comprises one or more exogenous polypeptides having homocitrate synthase, homocitrate dehydratase, homoaconitate hydratase, isohomocitrate dehydrogenase, 2-hydroxyglutarate dehydrogenase, glutaconate CoA-transferase, or 2-hydroxyglutaryl-CoA dehydratase activity;
or
the host further comprises one or more exogenous polypeptides having glutarate semialdehyde dehydrogenase, 4-hydroxy-2-oxoheptanedioate aldolase, 2-oxo-hept-3-ene-1,7-dioate hydratase, 2-enoate reductase, 2-hydroxyglutarate dehydrogenase, glutaconate CoA-transferase, or 2-hydroxyglutaryl-CoA dehydratase activity.

6 more elements

Dependencies are inferred from the claim text.

Methods, reagents and cells for biosynthesizing compound

Life of this patent

Methods, reagents and cells for biosynthesizing compound

Life of this patent

Abstract

Claims65