Genomic engineering of biosynthetic pathways leading to increased NADPH

Patent metadata

Field	Value
Publication number	US-11519012-B2
Application number	US-201816614566-A
Country	US
Kind code	B2
Filing date	May 18, 2018
Priority date	May 19, 2017
Publication date	Dec 6, 2022
Grant date	Dec 6, 2022

Abstract

The disclosure relates to host cells having altered NADPH availability, allowing for increased production of compounds produced using NADPH, and methods of use thereof. NADPH availability is altered by one or more of: expressing an altered GAPDH, expressing a variant glutamate dehydrogenase (gdh), aspartate semialdehyde dehydrogenase (asd), dihydropicolinate reductase (dapB), and meso-diaminopimelate dehydrogenase (ddh), expressing a novel nicotinamide nucleotide transhydrogenase, expressing a novel threonine aldolase, and expressing or modulating the expression of a pyruvate carboxylase in the host cells.

First claim

What is claimed is:

1. A method of improving a microbial cell's ability to produce L-lysine, the method comprising altering the microbial cell's available NADPH by genetically modifying the microbial cell to express:
a) an aldehyde dehydrogenase from Lactobacillus agilis comprising SEQ ID NO: 30 or an aspartate-semi-aldehyde dehydrogenase from C. glutamicum comprising SEQ ID NO:40;
b) an glutamate dehydrogenase from Clostridium symbiosum comprising SEQ ID NO: 44;
c) a 4-hydroxy-tetrahydrodipicolinate reductase from Escherichia coli comprising SEQ ID NO: 48; and
d) a meso-diaminopimelate D-dehydrogenase from C. glutamicum comprising SEQ ID NO: 4,

wherein the microbial cell produces at least 40% more L-Lysine than a microbial cell that does not contain a)-d).

2. The method of claim 1 , wherein the microbial cell is a microbial cell selected from the group consisting of Corynebacterium cell, Escherichia cell, Bacillus cell and Geobacillus cell.

3. The method of claim 1 , wherein the microbial cell is a Corynebacterium glutamicum cell or an Escherichia coli cell.

4. The method of claim 1 , wherein the bacteria is E. coli and the method further comprises expressing a pyruvate carboxylase (pyc) in the E. coli cell.

5. The method of claim 1 , wherein the microbial cell is a Corynebacterium cell.

6. The method of claim 1 , wherein the microbial cell is an Escherichia cell.

7. The method of claim 1 , herein the microbial cell is a Bacillus cell.

8. The method of claim 1 , wherein the microbial cell is a Geobacillus cell.

9. The method of claim 1 , wherein the method comprises altering the microbial cell's available NADPH by genetically modifying the microbial cell to express:
a) an aldehyde dehydrogenase Lactobacillus agilis comprising SEQ ID NO: 30;
b) an glutamate dehydrogenase from Clostridium symbiosum comprising SEQ ID NO: 44;
c) a 4-hydroxy-tetrahydrodipicolinate reductase from Escherichia coli comprising SEQ ID NO: 48; and
d) a meso-diaminopimelate D-dehydrogenase from C. glutamicum comprising SEQ ID NO: 4.

10. The method of claim 9 , wherein the microbial cell is a Corynebacterium cell.

11. The method of claim 1 , wherein the method comprises altering the microbial cell's available NADPH by genetically modifying the microbial cell to express:
a) an aspartate-semi-aldehyde dehydrogenase from C. glutamicum comprising SEQ ID NO:40;
b) an glutamate dehydrogenase from Clostridium symbiosum comprising SEQ ID NO: 44;
c) a 4-hydroxy-tetrahydrodipicolinate reductase from Escherichia coli comprising SEQ ID NO: 48; and
d) a meso-diaminopimelate D-dehydrogenase from C. glutamicum comprising SEQ ID NO: 4.

12. The method of claim 11 , wherein the microbial cell is a Corynebacterium cell.

Assignees

• Zymergen Inc

Inventors

• Manchester Shawn
• Mason Benjamin
• Goranov Alexi

Classifications

• C12P13/08 Primary

  Lysine; Diaminopimelic acid; Threonine; Valine · CPC title

• C12Y105/01017

  Alanopine dehydrogenase (1.5.1.17) · CPC title

• C12Y102/01012 Primary

  Glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) (1.2.1.12) · CPC title

• C12Y104/01003

  Glutamate dehydrogenase (NAD(P)+)(1.4.1.3) · CPC title

• C12Y117/01

  with NAD+ or NADP+ as acceptor (1.17.1) · CPC title