Genome-wide rationally-designed mutations leading to enhanced lysine production in E. coli

Patent metadata

Field	Value
Publication number	US-10920189-B2
Application number	US-202016904827-A
Country	US
Kind code	B2
Filing date	Jun 18, 2020
Priority date	Jun 21, 2019
Publication date	Feb 16, 2021
Grant date	Feb 16, 2021

Abstract

The present disclosure relates to various different types of variants in E. coli coding and noncoding regions leading to enhanced lysine production for, e.g., supplements and nutraceuticals.

First claim

We claim:

1. An engineered E. coil cell comprising the following variant sequences: a dapA protein having the amino add sequence of SEQ ID No. 1 and a dapA gene promoter sequence having the nucleic add sequence of SEQ ID No. 2 driving transcription of the dapA protein; and further comprising an additional variant protein selected from the following variant proteins: a lysC protein haying the amino add sequence of SEQ ID No. 3, a garb protein encoded by the nucleic add sequence of SEQ. ID No. 4, a yicL protein encoded by the nucleic add sequence of SEQ ID No. 5, a lysP protein having the amino add sequence of SEQ ID No. 6, a mgSA protein encoded by the nucleic add sequence of SEQ ID No. 7, and a pckE protein having the amino add sequence of SEQ ID No. 8.

2. The engineered E. coil cell of claim 1 , comprising the dapA protein having the amino acid sequence of SEQ ID No. 1, the dapA gene promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA protein, and further comprising the lysC protein having the amino acid sequence of SEQ ID No. 3.

3. The engineered E. coli cell of claim 1 , comprising the dapA protein having the amino acid sequence of SEQ ID No. 1 , the dapA gene promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA protein, and the garD protein coding sequence having the nucleic acid sequence of SEQ ID No. 4.

4. The engineered E. coli cell of claim 1 , comprising the dapA protein having the amino acid sequence of SEQ ID No. 1, the dapA gene promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA protein, and further comprising the yicL protein encoded by the nucleic add sequence of SEQ. ID No.5.

5. The engineered E coil cell of claim 1 , comprising the dapA protein having the amino acid sequence of SEQ. ID No. 1, the dapA gene promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA protein, and further comprising the lysP protein having the amino acid sequence of SEQ. ID No. 6.

6. The engineered E coil cell of claim 1 , comprising the dapA protein having the amino acid sequence of SEQ. ID No. 1, the dapA gene promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA protein, and further comprising the mgSA protein encoded by the nucleic add sequence SEQ. ID No. 7.

7. The engineered E. coil cell of claim 1 , comprising the dapA protein having the amino add sequence of SEQ ID No. 1, the dapA gene promoter sequence having the nucleic add SEQ ID No. 2, and further comprising the pckE protein having the amino add sequence of SEQ ID No. 8.

8. An engineered E. coil cell comprising the following variant sequences: a promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of a dapA gene, and further comprising one of the following proteins: an amyA protein encoded by the nucleic add sequence of SEQ ID No. 9, an amyA protein having the amino add sequence of SEQ ID No. 10, a cysN protein having the amino add sequence of SEQ ID No. 11, a dosP protein encoded by the nucleic add sequence of SEQ ID No. 12, a emrE protein encoded by the nucleic add sequence SEQ ID No. 13, a focB protein encoded by the nucleic add sequence SEQ ID No. 14, a glnD protein encoded by the nucleic add sequence SEQ. ID No. 15, a glnE protein having the amino acid sequence of SEQ ID No. 16, a hicB protein encoded by the nucleic add sequence of SEQ No. 17, a macB protein encoded by the nucleic acid sequence of SEQ ID No. 18, a marA protein having the amino add sequence of SEQ ID No. 19, or a metL protein having the amino add sequence of SEQ ID No. 20.

9. The engineered E coil cell of claim 8 comprising the promoter sequence haying the nucleic add SEQ ID No. 2 driving transcription of the dapA gene and further comprising the amyA protein coding sequence having the nucleic acid sequence of SEQ ID No. 9.

10. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and further comprising the amyA protein having the amino add sequence of SEQ ID No. 10.

11. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and further comprising the cysN protein having the amino acid sequence of SEQ ID No. 11.

12. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and the dosP protein encoded by the nucleic add sequence of SEQ No. 12.

13. The engineered E. coli cell of claim 8 comprising the promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA gene and further comprising the emrE protein encoded by the nucleic acid sequence of SEQ ID No. 13.

14. The engineered E. coli cell of claim 8 comprising the promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA gene and further comprising the focB protein encoded by the nucleic acid sequence of SEQ ID No. 14.

15. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2, driving transcription of the dapA gene and the glnD protein encoded by the nucleic add sequence of SEQ ID No. 15.

16. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and further comprising the glnE protein having the amino acid sequence of SEQ ID No. 16.

17. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and further comprising the hicB protein encoded by the nucleic acid sequence of SEQ ID No.17.

18. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA gene and further comprising the maeB protein encoded by the nucleic add sequence of SEQ ID No. 18.

19. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic add SEQ ID No. 2 driving transcription of the dapA gene and further comprising the marA protein having the amino acid sequence of SEQ ID No. 19.

20. The engineered E. coil cell of claim 8 comprising the promoter sequence having the nucleic acid SEQ ID No. 2 driving transcription of the dapA gene and further comprising the metL protein having the amino acid sequence of SEQ ID No. 20.

Assignees

• Inscripta Inc

Inventors

• Fox Richard
• Held Daniel
• Abbate Eric
• Clay Michael
• Krouse Katherine
• Krishnamurthy Nandini
• Yerramsetty Krishna

Classifications

• C12N2511/00

  Cells for large scale production · CPC title

• C12N9/88

  Lyases (4.) · CPC title

• C12N2510/02

  Cells for production · CPC title

• C12N1/20 Primary

  Bacteria; Culture media therefor · CPC title

• C07K14/245

  Escherichia (G) · CPC title