IP Library Granted Patent US 11,660,961
Granted Patent B2
US 11,660,961 · App. 17/067,610 · Granted May 30, 2023

Host cells and methods for producing isopentenol from mevalonate

Inventors: Taek Soon Lee (Berkeley, CA); Aram Kang (Richmond, CA)
Assignee: THE REGENTS OF THE UNIVERSITY OF CALIFORNIA
B60K35/00B65G69/006C07F9/091C07F9/098C08F136/08C12N9/0006C12N9/1025C12N9/1205C12N9/16C12N9/88C12P5/007C12P7/04C12Y101/01088C12Y203/0301C12Y207/01036C12Y401/01033B60K2370/173G01S2013/9314G01S2013/9317G01S2013/9323G01S2013/9324
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Quick Facts
Patent No.
US 11,660,961
App. No.
17/067,610
Granted
May 30, 2023
Kind
B2
Abstract

The present invention provides for a genetically modified host cell capable of producing isopentenol and/or 3-methyl-3-butenol, comprising (a) an increased expression of phosphomevalonate decarboxylase (PMD) (b) an increased expression of a phosphatase capable of converting isopentenol into 3-methyl-3-butenol, (c) optionally the genetically modified host cell does not express, or has a decreased expression of one or more of NudB, phosphomevalonate kinase (PMK), and/or PMD, and (d) optionally one or more further enzymes capable of converting isopentenol and/or 3-methyl-3-butenol into a third compound, such as isoprene.

Claims (25)

1. A polypeptide having a phosphomevalonate decarboxylase (PMD) enzymatic activity, and encoding an amino acid sequence comprising (a) at least 90% identity with SEQ ID NO:1, and (b) wherein (i) amino acid residue at position 74 is histidine, (ii) amino acid residue at position 145 is phenylalanine, or (iii) amino acid residue at position 74 is histidine and amino acid residue at position 145 is phenylalanine, corresponding to the numbering of SEQ ID NO:1.

2. The polypeptide of claim 1 , wherein the amino acid sequence comprises (i) a histidine at position 74, (ii) a phenylalanine at position 145, or (iii) a histidine at position 74 and a phenylalanine at position 145.

3. The polypeptide of claim 1 , wherein the amino acid sequence comprises the following amino acid residues: E at position 71, S at position 108, N at position 110, A at position 119, S at position 120, S at position 121, A at position 122, S at position 155, Rat position 158, S at position 208, and D at position 302 corresponding to SEQ ID NO:1.

4. The polypeptide of claim 1 , wherein the amino acid sequence comprises at least 95% identity with SEQ ID NO:1.

5. The polypeptide of claim 4 , wherein the amino acid sequence comprises at least 99% identity with SEQ ID NO:1.

6. A vector encoding the polypeptide of claim 1 , and which is operatively linked to a promoter.

7. A genetically modified host cell comprising the vector of claim 6 , wherein the genetically modified host cell is capable of expressing the polypeptide, and the genetically modified host cell is a bacterial or fungal cell.

8. The genetically modified host cell of claim 7 , wherein the genetically modified host cell is a species of the genus Escherichia, Enterobacter, Azotobacter, Erwinia, Bacillus, Pseudomonas, Klebsielia, Proteus, Salmonella, Serratia, Shigella, Rhizobia, Vitreoscilla, Paracoccus , or Clostridia.

9. The genetically modified host cell of claim 8 , wherein the genetically modified host cell is a species of the genus Escherichia.

10. The genetically modified host cell of claim 9 , wherein the genetically modified host cell is Escherichia coli.

11. The genetically modified host cell of claim 7 , wherein the genetically modified host cell is a yeast cell.

12. The genetically modified host cell of claim 11 , wherein the yeast cell is a species of the Saccharomyces genus.

13. The genetically modified host cell of claim 12 , wherein the yeast cell is Saccharomyces cerevisiae.

14. A method for producing 3-methyl-3-butenol, comprising:

(a) providing a genetically modified host cell of claim 9 wherein the genetically modified host cell is capable of producing mevalonate; and

(b) culturing the genetically modified host cell under a condition wherein the polypeptide is expressed and 3-methyl-3-butenol is produced.

15. The method of claim 14 , wherein the genetically modified host cell does not express, or has a decreased expression of phosphomevalonate kinase (PMK) and/or NudB phosphatase (NudB).

16. The method of claim 15 , wherein the genetically modified host cell does not express, or has a decreased expression of PMK.

17. The method of claim 15 , wherein the genetically modified host cell does not express, or has a decreased expression of NudB.

18. The method of claim 14 , wherein the (b) culturing step is under an anaerobic or microaerobic condition.

19. The method of claim 14 , comprising (c) recovering the 3-methyl-3-butenol.

20. The method of claim 14 , wherein the genetically modified host cell comprises one or more further enzymes capable of converting 3-methyl-3-butenol into a third compound.

21. The method of claim 20 , comprising (c) recovering the third compound.

22. The method of claim 21 , wherein the third compound is isoprene.

23. The method of claim 22 , comprising (c) recovering the isoprene.

Assignments (2)
CONFIRMATORY LICENSE Recorded Feb 22, 2021
From: UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
To: UNITED STATES DEPARTMENT OF ENERGY
Reel/Frame 055353/0968 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Oct 13, 2020
From: LEE, TAEK SOON; KANG, ARAM
To: THE REGENTS OF THE UNIVERSITY OF CALIFORNIA
Reel/Frame 054041/0641 →
Continuity (5)
Continuation 16388556 · Apr 18, 2019
Continuation 15682325 · Aug 21, 2017
Continuation PCTUS2016018984 · Feb 22, 2016
Provisional Application 62119071 · Feb 20, 2015
Related Publication 20210023947A1 · Jan 28, 2021