Methods of producing 7-carbon chemicals via methyl-ester shielded carbon chain elongation

Patent metadata

Field	Value
Publication number	US-10196657-B2
Application number	US-201314139072-A
Country	US
Kind code	B2
Filing date	Dec 23, 2013
Priority date	Dec 31, 2012
Publication date	Feb 5, 2019
Grant date	Feb 5, 2019

Abstract

This document describes biochemical pathways for producing pimelic acid, 7-aminoheptanoic acid, 7-hydroxyheptanoic acid, heptamethylenediamine or 1,7-heptanediol by forming two terminal functional groups, comprised of carboxyl, amine or hydroxyl group, in a C7 aliphatic backbone substrate. These pathways, metabolic engineering and cultivation strategies described herein rely on enzymes or homologs accepting methyl ester shielded dicarboxylic acid substrates.

First claim

What is claimed is:

1. A method for biosynthesizing pimelic acid or 7-aminoheptanoate, said method comprising:
(i) enzymatically converting malonyl-[acp] to pimeloyl-[acp] via (1) contacting malonyl-[acp] with a polypeptide having the activity of a malonyl-[acp] O-methyltransferase classified under EC 2.1.1.197 to form malony-[acp] methyl ester, (2) contacting malonyl-[acp] methyl ester and malony-[acp] with a polypeptide having the activity of a β-ketoacyl-[acp] synthase classified under EC 2.3.1.- to form 3-oxoglutyryl-[acp] methyl ester, (3) contacting 3-oxoglutyryl-[acp] methyl ester with a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified under EC 1.1.1.100 to form 3-hydroxy-glutaryl-[acp] methyl ester, (4) contacting 3-hydroxy-glutaryl-[acp] methyl ester with a polypeptide having the activity of a 3-hydroxyacyl-[acp] dehydratase classified under EC 4.2.1.59 to form 2,3-dehydroglutaryl-[acp] methyl ester, (5) contacting 2,3-dehydroglutaryl-[acp] methyl ester with a polypeptide having the activity of an enoyl-[acp] reductase classified under EC 1.3.1.10 to form glutaryl-[acp] methyl ester, (6) contacting glutaryl-[acp] methyl ester with a polypeptide having the activity of a β-ketoacyl-[acp] synthase classified under EC 2.3.1.- to form 3-oxopimeloyl-[acp] methyl ester, (7) contacting 3-oxopimeloyl-[acp] methyl ester with a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified under EC 1.1.1.100 to form 3-hydroxy-pimeloyl-[acp] methyl ester, (8) contacting 3-hydroxy-pimeloyl-[acp] methyl ester with a polypeptide having the activity of a 3-hydroxyacyl-[acp] dehydratase classified under EC 4.2.1.59 to form 2,3-dehydropimeloyl-[acp] methyl ester, (9) contacting 2,3-dehydropimeloyl-[acp] methyl ester with a polypeptide having the activity of an enoyl-[acp] reductase classified under EC 1.3.1.10 to form pimeloyl-[acp] methyl ester, and (10) contacting pimeloyl-[acp] methyl ester with a polypeptide having the activity of a pimelyl-[acp] methyl ester esterase classified under EC 3.1.1.85 to form pimeloyl-[acp];
enzymatically converting malonyl-CoA to pimeloyl-CoA via (1) contacting malonyl-CoA with a polypeptide having the activity of a malonyl-[acp] O-methyltransferase classified under EC 2.1.1.197 to form malonyl-CoA methyl ester, (2) contacting malonyl-CoA methyl ester and acetyl-CoA with a polypeptide having the activity of a β-ketathiolase classified under EC 2.3.1.16 or a β-ketoacyl-[acp] synthase classified under EC 2.3.1.180 to form 3-oxoglutaryl-CoA methyl ester, (3) contacting 3-oxoglutaryl-CoA methyl ester with a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified under EC 1.1.1.100 or an acetoacetyl-CoA reductase classified under EC 1.1.1.36 to form 3-hydroxy-glutaryl-CoA methyl ester, (4) contacting 3-hydroxy-glutaryl-CoA methyl ester with a polypeptide having the activity of an enoyl-CoA hydratase classified under EC 4.2.1.119 to form 2,3-dehydroglutaryl-CoA methyl ester, (5) contacting 2,3-dehydroglutaryl-CoA methyl ester with a polypeptide having the activity of an enoyl-[acp] reductase classified under EC 1.3.1.10 or a trans-2-enoyl-CoA reductase classified under EC 1.3.1.38, EC 1.3.1.8, or EC 1.3.1.44 to form glutaryl-CoA methyl ester, (6) contacting glutaryl-CoA methyl ester with a polypeptide having the activity of a β-ketoacyl-[acp] synthase classified under EC 2.3.1.- or a β-ketothiolase classified under EC 2.3.1.16 to form 3-oxopimeloyl-CoA methyl ester, (7) contacting 3-oxopimeloyl-CoA methyl ester with a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified under EC 1.1.1.100 or an acetoacetyl-CoA reductase classified under EC 1.1.1.36 to form 3-hydroxy-pimeloyl-CoA methyl ester, (8) contacting 3-hydroxy-pimeloyl-CoA methyl ester with a polypeptide having the activity of an enoyl-CoA hydratase classified under EC 4.2.1.119 to form 2,3-dehydropimeloyl-CoA methyl ester, (9) contacting 2,3-dehydropimeloyl-CoA methyl ester with a polypeptide having the activity of an enoyl-[acp] reductase classified under EC 1.3.1.10 or a trans-2-enoyl-CoA reductase classified under EC 1.3.1.38, EC 1.3.1.8, or EC 1.3.1.44 to form pimeloyl-CoA methyl ester, and (10) contacting pimeloyl-CoA methyl ester with a polypeptide having the activity of a pimelyl-[acp] methyl ester esterase classified under EC 3.1.1.85 to form pimeloyl-CoA; or
enzymatically converting malonyl-CoA to pimeloyl-CoA via (1) contacting malonyl-CoA with a polypeptide having the activity of a malonyl-[acp] O-methyltransferase classified under EC 2.1.1.197 to form malonyl-CoA methyl ester, (2) contacting malonyl-CoA methyl ester and acetyl-CoA with a polypeptide having the activity of a β-ketothiolase classified under EC 2.3.1.16 or a β-ketoacyl-[acp] synthase classified under EC 2.3.1.180 to form 3-oxoglutaryl-CoA methyl ester, (3) contacting 3-oxoglutaryl-CoA methyl ester with a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified under EC 1.1.1.35 or EC 1.1.1.157 to form 3-hydroxy-glutaryl-CoA methyl ester, (4) contacting 3-hydroxy-glutaryl-CoA methyl ester with a polypeptide having the activity of an enoyl-CoA hydratase classified under EC 4.2.1.17 to form 2,3-dehydroglutaryl-CoA methyl ester, (5) contacting 2,3-dehydroglutaryl-CoA methyl ester with a polypeptide having the activity of a trans-2-enoyl-CoA reductase classified under EC 1.3.1.44 to form glutaryl-CoA methyl ester, (6) contacting glutaryl-CoA methyl ester with a polypeptide having the activity of a β-ketoacyl-[acp] synthase classified under EC 2.3.1.- or a β-ketathiolase classified under EC 2.3.1.16 to 3-oxopimeloyl-CoA methyl ester, (7) contacting 3-oxopimeloyl-CoA methyl ester with a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified under EC 1.1.1.35 or EC 1.1.1.157 to form 3-hydroxy-pimeloyl-CoA methyl ester, (8) contacting 3-hydroxy-pimeloyl-CoA methyl ester with a polypeptide having the activity of an enoyl-CoA hydratase classified under EC 4.2.1.17 to form 2,3-dehydropimeloyl-CoA methyl ester, (9) contacting 2,3-dehydropimeloyl-CoA methyl ester with a polypeptide having the activity of a trans-2-enoyl-CoA reductase classified under EC 1.3.1.44 to form pimeloyl-CoA methyl ester, and (10) contacting pimeloyl-CoA methyl ester with a polypeptide having the activity of a pimelyl-[acp] methyl ester esterase classified under EC 3.1.1.85 to form pimeloyl-CoA; and
(ii) (a) enzymatically converting pimeloyl-CoA or pimeloyl-[acp] to pimelic acid via (1) contacting pimeloyl-CoA or pimeloyl-[acp] with a polypeptide having the activity of a thioesterase classified under EC 3.1.2.- to form pimelic acid, (2) contacting pimeloyl-CoA with a polypeptide having the activity of a glutaconate CoA-transferase classified under EC 2.8.3.12 or a reversible succinate-CoA ligase classified under EC 6.2.1.5 to form pimelic acid, or (3) contacting pimeloyl-CoA with a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified under EC 1.2.1.10 to form pimelate semialdehyde and contacting pimelate semialdehyde with a polypeptide having the activity of a 7-oxoheptanoate dehydrogenase classified under EC 1.2.1.-, a 6-oxohexanoate dehydrogenase classified under EC 1.2.1.-, or an aldehyde dehydrogenase classified under EC 1.2.1.3 to form pimelic acid; or
(b) enzymatically converting pimeloyl-CoA or pimeloyl-[acp] to 7-aminoheptanoate via (1) contacting pimeloyl-CoA with a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified under EC 1.2.1.10 to form pimelate semialdehyde and contacting pimelate semialdehyde with a polypeptide having the activity of a ω-transaminase classified under EC 2.6.1.- to form 7-aminoheptanoate, or (2) contacting any one of pimelic acid formed in (ii)(a) with a polypeptide having the activity of a carboxylate reductase classified under EC 1.2.99.6 to form pimelate semialdehyde and contacting pimelate semi

Assignees

• Invista North America Sarl
• Invista North America Sarl

Inventors

• Pearlman Paul S
• Conradie Alex Van Eck
• Chen Changlin
• Botes Adriana Leonora

Classifications

• C12N9/1007

  Methyltransferases (general) (2.1.1.) · CPC title

• C12N9/88

  Lyases (4.) · CPC title

• C12P13/001 Primary

  Amines; Imines · CPC title

• C12P7/44

  Polycarboxylic acids · CPC title

• C12N9/0006

  acting on CH-OH groups as donors (1.1) · CPC title