Production of lysergic acid by genetic modification of a fungus

Patent metadata

Field	Value
Publication number	US-9790528-B2
Application number	US-201514739382-A
Country	US
Kind code	B2
Filing date	Jun 15, 2015
Priority date	Jun 16, 2014
Publication date	Oct 17, 2017
Grant date	Oct 17, 2017

Abstract

The present invention provides a method of producing lysergic acid and other ergot alkaloids by genetic modification of a fungus. A strain of fungus comprising Aspergillus fumigatus ( A. fumigatus ) and expressing one or more genes of the ergot alkaloid biosynthesis pathway from one or more fungus selected from the group consisting of Epichloë festucae var. lolii×Epichloë typhina isolate Lp1 ( E . sp. Lp1); Claviceps species; Claviceps africana ( C. africana ); Claviceps gigantea ( C. gigantea ); Epichloë coenophiala and Periglandula species, wherein gene easA or gene easM is inactivated in said A. fumigatus , is provided.

First claim

What is claimed is:

1. A method for producing lysergic acid comprising inactivating an ergot alkaloid biosynthesis pathway gene from the fungus Aspergillus fumigatus and expressing genes easA and cloA from the fungus Epichloe sp. Lp1, wherein said inactivated ergot alkaloid biosynthesis pathway gene is easA of Aspergillus fumigatus.

2. A method for producing ergot alkaloids comprising inactivating an ergot alkaloid biosynthesis pathway gene from the fungus A. fumigatus and expressing genes easA and cloA from the fungus E. sp. Lp1, wherein said inactivated ergot alkaloid biosynthesis pathway gene is easA of A. fumigants.

3. A method for producing dihydrolysergic acid (DHLA) comprising inactivating gene easM in A. fumigatus and expressing gene cloA from Claviceps gigantea or gene cloA from Claviceps africana in said A. fumigatus strain.

4. A method for producing dihydrolysergol (DHlysergol) comprising inactivating gene easM in A. fumigatus and expressing one or more genes of the ergot alkaloid biosynthesis from Claviceps gigantea selected from the group consisting of:
a. cloA; and
b. cloA and easA,
wherein said gene(s) from C. gigantea are expressed in said A. fumigants strain.

5. A method of producing ergot alkaloids in a strain of A. fumigatus comprising expressing a bidirectional easA/easG promoter of A. fumigatus to drive expression of cloA oxidase genes in the A. fumigatus EasA knock-out background.

6. The method of claim 5 , wherein a EasA gene from E. sp. Lp1 is expressed in said A. fumigatus EasA knock-out background.

7. The method of claim 6 , wherein said EasA gene from E. sp. Lp1 includes expression of cloA.

8. A method for the production of lysergic acid comprising providing for the expression of ergot alkaloid producing fungi EasA/CloA in A. fumigants easA knockout or easM knockout.

9. A method for producing lysergic acid in A. fumigatus easA knock-out providing amplifying E. sp. Lp1 easA and E. sp. Lp1 cloA for producing lysergic acid.

10. A method for accumulating lysergol comprising amplifying easA and cloA from Periglandula in plant material selected from the group consisting of Stictocardia tiliifolia, S. beraviensis, Argyreia , and Ipomoea species or by amplifying easA and cloA from Epichloë coenophiala for accumulating lysergol.

11. A method for producing lysergol comprising providing expressing Periglandula sp. easA and P. sp. cloA or Epichloë coenophiala easA and Epichloë coenophiala cloA in a A. fumigants easA knock-out strain for producing lysergol.

12. A method for producing lysergol comprising A. fumigatus easA knock-out strain through expression of E. sp. Lp1 easA and P. sp. cloA or Epichloë coenophiala cloA for producing lysergol.

13. The method according to claim 12 , including amplifying said easA and cloA based on degenerate primers designed to anneal to versions of each gene.

14. A method for producing dihydrolysergic acid comprising expressing C. africana cloA under the control of a A. fumigatus easA promoter in A. fumigants easM knock-out strain for producing dihydrolysergic acid.

15. A method for producing dihydrolysergic acid comprising expressing C. africana easA, C. africana cloA, and E . sp. Lp1 cloA using a A. fumigatus easA/easG promoter in a A. fumigatus easM knock-out strain for producing dihydrolysergic acid.

16. A method for producing dihydrolysergol comprising expressing C. gigantea cloA in A. fumigatus easM knock-out for producing dihydrolysergol.

17. The method of claim 16 , including joining said cloA to a easA promoter of A. fumigatus to form a cloA construct and introducing said cloA construct into A. fumigants easM knock-out utilizing cotransformation with pBCphleo.

18. The method of claim 17 , including adding a C. gigantea easA operably linked to a A. fumigants easA promoter to said cotransformation of the A. fumigatus easM knockout.

Assignees

• Univ West Virginia

Inventors

• Panaccione Daniel G
• Robinson Sarah L

Classifications

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title

• C12P17/182 Primary

  Heterocyclic compounds containing nitrogen atoms as the only ring heteroatoms in the condensed system (alloxazine or isoalloxazine, e.g. riboflavine C12P25/00) · CPC title

• C12N1/14

  Fungi (culture of mushrooms A01G18/00; as new plants A01H15/00); Culture media therefor · CPC title