# US 10,246,688 B2 · Diterpenoid synthesis

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 10,246,688 B2
- Document ID: US10246688B2
- Status: Granted
- Kind: B2
- Application number: 15/105,502
- Filed: 2015-01-09
- Published: 2016-11-03
- Granted: 2019-04-02
- Earliest claimed: 2014-01-13
- Assignees: The University of York
- Applicants: The University of York
- Inventors: Andrew King; Ian Graham
- Web page: https://patentgrep.com/patent/US10246688B2

## CPC

- C12N 9/0071: {acting on paired donors with incorporation of molecular oxygen (1.14)}
- C12N 15/52: Genes encoding for enzymes or proenzymes
- C12N 15/8243: {involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine}
- C12P 7/02: containing a hydroxy group
- C12P 7/26: Ketones
- 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)}

## Abstract

The present disclosure relates to nucleic acids that encode polypeptides with cytochrome P450 activity involved in the biosynthesis of plant derived diterpenoids or diterpenes.

## Claims

### Claim 1

A transgenic fungal cell comprising an expression vector adapted to express a nucleic acid molecule, wherein the nucleic acid molecule comprises:
i) the nucleotide sequence of SEQ ID NO: 6;
ii) a nucleotide sequence comprising at least 90% identity to SEQ ID NO: 6 and that encodes a polypeptide having cytochrome P450 activity; or
iii) a nucleotide sequence degenerate as a result of the genetic code to the nucleotide sequence of SEQ ID NO: 6 and encodes a polypeptide comprising SEQ ID NO: 32 and that has cytochrome P450 activity;
or wherein the nucleic acid molecule comprises:
iv) the nucleotide sequence of SEQ ID NO: 18;
v) a nucleotide sequence comprising at least 90% identity to SEQ ID NO: 18 and that encodes a polypeptide having cytochrome P450 activity; or
vi) a nucleotide sequence degenerate as a result of the genetic code to the nucleotide sequence of SEQ ID NO: 18 and encodes the polypeptide of SEQ ID NO: 44 and that has cytochrome P450 activity.

### Claim 2 (depends on 1)

A process for modification of one or more diterpenoids or diterpenes, the process comprising:
i) cultivating the transgenic fungal cell of claim 1; and optionally
ii) isolating said one or more diterpenoid or diterpene from the transgenic fungus or fungal culture.

### Claim 3 (depends on 1)

The transgenic fungal cell of claim 1, wherein said fungal cell is a Saccharomyces cerevisiae cell.

### Claim 4 (depends on 3)

A process for modification of one or more diterpenes or diterpenoids, the process comprising:
i) cultivating the transgenic Saccharomyces cerevisiae cell of claim 3 under conditions that modify one or more diterpenes or diterpenoids; and optionally
ii) isolating said one or more diterpenoid or diterpene from the transgenic Saccharomyces cerevisiae cell or cell culture.

### Claim 5 (depends on 1)

The transgenic fungal cell of claim 1, wherein the fungal cell is transformed or transfected with an expression vector adapted to express the nucleic acid molecule of SEQ ID NO: 6.

### Claim 6 (depends on 1)

The transgenic fungal cell according to claim 1, wherein the fungal cell is transformed or transfected with an expression vector adapted to express the nucleic acid molecule of SEQ ID NO: 18.

### Claim 7 (depends on 1)

The transgenic fungal cell of claim 1, wherein the fungal cell is transformed or transfected with an expression vector adapted to express the nucleotide sequence degenerate as a result of the genetic code to the nucleotide sequence of SEQ ID NO: 6 and encodes the amino acid sequence of SEQ ID NO: 32.

### Claim 8 (depends on 1)

The transgenic fungal cell of claim 1, wherein the fungal cell is transformed or transfected with an expression vector adapted to express the nucleotide sequence degenerate as a result of the genetic code to the nucleotide sequence of SEQ ID NO: 18 and encodes the amino acid sequence of SEQ ID NO: 44.

### Claim 9 (depends on 1)

The transgenic fungal cell of claim 1, wherein the nucleic acid molecule expressed by the expression vector is over-expressed when compared to a non-transgenic fungal cell of the same species.