Auto-inducible expression system

Patent metadata

Field	Value
Publication number	US-10106801-B2
Application number	US-201515514028-A
Country	US
Kind code	B2
Filing date	Sep 21, 2015
Priority date	Sep 24, 2014
Publication date	Oct 23, 2018
Grant date	Oct 23, 2018

Abstract

A method for the expression of a protein of interest by a bacterium, notable in that it comprises the culturing of a bacterium temporarily or continuously expressing an Hsp protein, in that said bacterium also comprises a nucleic acid sequence, encoding a protein of interest, under the control of a lac promoter and in that said bacterium is cultured in a medium which does not contain IPTG or a metabolized molecule in such a way as to automatically induce the induction of transcription from the lac promoter.

First claim

The invention claimed is:

1. A method for an expression of a protein of interest by a bacterium comprising:
culturing of the bacterium temporarily or continuously expressing an Hsp protein encoded by a first nucleic acid sequence under a control of a lac promoter,
wherein said bacterium further comprises a second nucleic acid sequence encoding a protein of interest under the control of the lac promoter, and
wherein said bacterium is cultured in a medium not containing IPTG or a metabolised molecule in such a way as to automatically induce an induction of transcription from the lac promoter.

2. The method according to claim 1 , wherein said second nucleic acid encoding the protein of interest is comprised in a first plasmid.

3. The method according claim 1 , wherein said protein of interest is a recombinant protein.

4. The method according to claim 1 , wherein said protein of interest is a polymerase.

5. The method according to claim 4 , wherein said polymerase is a T7 polymerase.

6. The method according to claim 4 , wherein said bacterium further comprises a third nucleic acid sequence encoding a second protein of interest placed under the control of a specific promoter of said polymerase.

7. The method according to claim 6 , wherein said third nucleic acid sequence is comprised in a first plasmid or in a second plasmid.

8. The method according to claim 1 , wherein said bacterium is an E. coli.

9. The method according to claim 8 , wherein said bacterium belongs to a BL21(DE3) strain or to a BL21(DE3) star strain.

10. The method according to claim 1 , wherein the Hsp protein is chosen in the group comprising Hsp40, Hsp70, Hsp90 and Hsp110.

11. The method according to claim 1 , wherein the Hsp protein is human Hsp70 protein.

12. The method according to claim 1 , wherein said bacterium is cultured in a liquid medium.

13. A method for an expression of a protein of interest by a bacterium comprising:
culturing of the bacterium temporarily or continuously expressing a human Hsp70 protein encoded by a first nucleic acid sequence,
wherein said bacterium further comprises a second nucleic acid sequence encoding a protein of interest under a control of a lac promoter, and
wherein said bacterium is cultured in a medium not containing IPTG or a metabolised molecule in such a way as to automatically induce an induction of transcription from the lac promoter.

14. The method according to claim 13 , wherein said second nucleic acid encoding the protein of interest is comprised in a first plasmid.

15. The method according to claim 13 , wherein said protein of interest is a recombinant protein.

16. The method according to claim 13 , wherein said protein of interest is a polymerase.

17. The method according to claim 16 , wherein said polymerase is a T7 polymerase.

18. The method according to claim 16 , wherein said bacterium further comprises a third nucleic acid sequence encoding a second protein of interest placed under the control of a specific promoter of said polymerase.

19. The method according to claim 18 , wherein said third nucleic acid sequence is comprised in a first plasmid or in a second plasmid.

20. The method according to claim 13 , wherein said human Hsp70 protein is encoded by the first nucleic acid sequence under the control of the lac promoter.

21. The method according to claim 13 , wherein said bacterium is an E. coli.

22. The method according to claim 21 , wherein said bacterium belongs to a BL21(DE3) strain or to a BL21(DE3) star strain.

23. The method according to claim 13 , wherein said bacterium is cultured in a liquid medium.

Assignees

• Univ Bourgogne
• Inra
• Inst Nat Sante Rech Med

Inventors

• Neiers Fabrice
• Seigneuric Renaud
• Briand Loïc
• Garrido-Fleury Carmen

Classifications

• C12N9/1247

  DNA-directed RNA polymerase (2.7.7.6) · CPC title

• C12N15/74

  Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora · CPC title

• C12N1/20

  Bacteria; Culture media therefor · CPC title

• C12N15/72 Primary

  Expression systems using regulatory sequences derived from the lac-operon · CPC title

• C12P21/02

  having a known sequence of two or more amino acids, e.g. glutathione · CPC title