# US 10,119,126 B2 · Enzymes, enzyme components and uses thereof

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- Number: US 10,119,126 B2
- Document ID: US10119126B2
- Status: Granted
- Kind: B2
- Application number: 15/864,798
- Filed: 2018-01-08
- Published: 2018-05-24
- Granted: 2018-11-06
- Earliest claimed: 2012-08-03
- Assignees: BASF Plant Science GmbH
- Applicants: BASF Plant Science Company GmbH
- Inventors: Laurent Marty; Toralf Senger
- Web page: https://patentgrep.com/patent/US10119126B2

## CPC

- C12N 9/0071: {acting on paired donors with incorporation of molecular oxygen (1.14)}
- C12N 9/001: {acting on the CH-CH group of donors (1.3)}
- C12P 7/6427: Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
- C12P 7/6432: Eicosapentaenoic acids [EPA]
- C12P 7/6434: Docosahexenoic acids [DHA]
- C12Y 114/19: with oxidation of a pair of donors resulting in the reduction of molecular oxygen to two molecules of water (1.14.19)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}
- C12Y: ENZYMES

## Abstract

The invention provides nucleic acid molecules which encodes a novel fatty acid desaturase, KCS, KCR and/or LACS from Thraustochytrium aureum and Sphaeroforma arctica. The invention also provides recombinant expression vectors containing the nucleic acid molecules, host cells into which the expression vectors have been introduced, and methods for large-scale production of long chain polyunsaturated fatty acids (LCPUFAs), e.g., ARA, EPA and DHA and for screening for delta-4 desaturases.

## Claims

### Claim 1

A method for increasing the efficiency of eicosapentaenoic acid (EPA) production in a host cell capable of synthesizing EPA, the method comprising expressing in the cell a heterologous lysophospholipid-coenzyme A synthase (LACS) enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity.

### Claim 2 (depends on 1)

The method of claim 1, wherein a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase is/are expressed in the cell.

### Claim 3

A plant comprising a host cell capable of synthesizing EPA, the host cell comprising a polynucleotide encoding a heterologous LACS enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity and further comprising one or more polynucleotides encoding a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase.

### Claim 4

A seed comprising a host cell capable of synthesizing EPA, the host cell comprising a polynucleotide encoding a heterologous LACS enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity and further comprising one or more polynucleotides encoding a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase.

### Claim 5

A method for increasing the efficiency of docosahexaenoic acid (DHA) production in a host cell capable of synthesizing DHA, the method comprising expressing in the cell a heterologous lysophospholipid-coenzyme A synthase (LACS) enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity.

### Claim 6 (depends on 5)

The method of claim 5, wherein a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase is/are expressed in the cell.

### Claim 7

A plant comprising a host cell capable of synthesizing DHA, the host cell comprising a polynucleotide encoding a heterologous LACS enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity and further comprising one or more polynucleotides encoding a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase.

### Claim 8

A seed comprising a host cell capable of synthesizing DHA, the host cell comprising a polynucleotide encoding a heterologous LACS enzyme having at least 80% sequence identity to SEQ ID NO: 84 and fatty acyl CoA synthetase activity and further comprising one or more polynucleotides encoding a delta-5 elongase, a delta-6 elongase, a delta-5 desaturase, and/or a delta-6 desaturase.