IP Library › Granted Patent US 12,203,078
Granted Patent B2
US 12,203,078 · App. 16/612,042 · Granted Jan 21, 2025

Inducible promoter for gene expression and synthetic biology

Inventors: Jean-Marc Nicaud (Trappes, FR); Marion Trassaert (Hazebrouck, FR); Stephane Thomas (Bois d'Arcy, FR); Patrick Fickers (Liege, BE); Marie Vandermies (Ottignies, BE); Frederic Carly (Brussels, BE)
Assignees: INSTITUT NATIONAL DE RECHERCHE POUR L'AGRICULTURE, L'ALIMENTATION ET L'ENVIRONNEMENT; UNIVERSITE LIBRE DE BRUXELLES; UNIVERSITE DE LIEGE
C12N15/815C07K14/39C12N9/1205C12N2310/51C12N2830/34C12Y101/01009C12Y207/01027
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Quick Facts
Patent No.
US 12,203,078
App. No.
16/612,042
Granted
Jan 21, 2025
Kind
B2
Abstract

The invention is related to an inducible promoter for improved and regulated gene expression, useful in synthetic biology and metabolic engineering. In particular, the present invention relates to a nucleotide sequence comprising the regulatory regions of an erythritol- and erythrulose-inducible promoter in yeast and uses thereof in an expression system thus allowing an improved and regulated gene expression and production of gene product.

Claims (39)

1. A method for expressing a gene or producing an encoded protein, comprising the steps of

a) growing in vitro, in a culture medium, a host cell comprising

an erythritol-inducible promoter sequence, an erythrulose-inducible promoter sequence or an erythritol- and erythrulose-inducible promoter sequence functional in yeast comprising a core promoter, and a nucleotide sequence comprising tandem repeats of a sequence according to formula (I) and/or tandem repeats of a sequence according to formula (II), wherein:

formula (I) is

CGGNNX 1 CNNNANNX 2 GNNAAGNCG (I)

with N being any nucleotide

with X 1 and X 2 being any nucleotide or no nucleotide,

formula (II) is

ANTTNNNTTTCCNNATNNGG (II)

with N being any nucleotide,

operably linked to a polynucleotide sequence encoding a gene product to be transcribed from said inducible promoter and

b) adding erythritol and/or erythrulose to the culture medium.

2. The method according to claim 1 , wherein the sequence according to formula (I) is selected in the group consisting of:

(SEQ ID NO: 6)

CGGVWYCYBVAWDGRRAAGSCG ,

(SEQ ID NO: 7)

CGGVWCYBVAWDKGRRAAGSCG ,

(SEQ ID NO: 8)

CGGVWYCYBVAWDKGRRAAGSCG ,

and

(SEQ ID NO: 9)

CGGVWCYBVAWDGRRAAGSCG .

3. The method according to claim 1 , wherein the sequence according to formula (I) is selected in the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 9.

4. The method according to claim 1 , wherein the sequence according to formula (II) is ABTTSYRTTTCCYWATDHGG (SEQ ID NO: 10).

5. The method according to claim 1 , wherein said nucleotide sequence contains from 2 to 32 copies of the sequence according to formula (I) and/or the sequence according to formula (II).

6. The method according to claim 1 , wherein said nucleotide sequence comprises at least one sequence according to formula (I) and further comprises at least one sequence according to formula (III) which is

CNTGCATNATCCGANGAC  (III)

with N being any nucleotide.

7. The method according to claim 6 , wherein the sequence according to formula (III) is CDTGCATWATCCGAYGAC as set forth in SEQ ID NO: 12.

8. The method according to claim 1 , wherein said nucleotide sequence comprises the sequence as set forth in SEQ ID NO: 13.

9. The method according to claim 1 wherein at least one sequence according to formula (I) and/or at least one sequence according to formula (II) is/are upstream to an ATG of the polynucleotide sequence.

10. The method according to claim 9 , wherein said at least one sequence according to formula (I) and/or said at least one sequence according to formula (II) is/are from −800 to −50 of the ATG.

11. The method of claim 1 wherein said core promoter is selected from the group consisting of TEF core promoter, EYK1 core promoter, EYD1 core promoter, POX2 core promoter, LEU2 core promoter and PAT1 core promoter.

12. The method of claim 1 , wherein said erythritol-inducible promoter sequence, erythrulose-inducible promoter sequence, or erythritol and erythrulose-inducible promoter sequence comprises at least one promoter regulatory sequence selected from the group consisting of UAS, TATA box and URS.

13. The method of claim 1 , wherein an endogenous gene encoding L-erythrulose kinase is inhibited in the host cell.

14. The method of claim 1 , an endogenous gene encoding erythritol dehydrogenase is inhibited in the host cell.

15. The method of claim 12 , wherein said promoter regulatory sequence is a regulatory sequence of a promoter selected from the group consisting of XPR2, TEF, POX2, LEU2, PAT1 and LIP2.

16. The method of claim 1 , wherein an endogenous gene encoding L-erythrulose kinase is deleted in the host cell.

17. The method of claim 1 , wherein an endogenous gene encoding erythritol dehydrogenase is deleted in the host cell.

Assignments (2)
CHANGE OF NAME Recorded Jun 15, 2021
From: INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE (INRA)
To: INSTITUT NATIONAL DE RECHERCHE POUR L'AGRICULTURE, L'ALIMENTATION ET L'ENVIRONNEMENT
Reel/Frame 056592/0475 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 20, 2021
From: NICAUD, JEAN-MARC; TRASSAERT, MARION; THOMAS, STEPHANE; FICKERS, PATRICK; VANDERMIES, MARIE; CARLY, FREDERIC
To: INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE; UNIVERSITE LIBRE DE BRUXELLES; UNIVERSITE DE LIEGE
Reel/Frame 054967/0643 →
Priority Claims (1)
EP 17305575 · May 18, 2017 · regional
Continuity (1)
Related Publication 20200149055A1 · May 14, 2020
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