Recombinant microorganism having ability to produce [lactate-co-glycolate] copolymer from glucose, and method for preparing [lactate-co-glycolate] copolymer using same

• US 8,883,463 B2
• Filed: 03/09/2012
• Issued: 11/11/2014
• Est. Priority Date: 03/11/2011
• Status: Active Grant

• Current Assignee: Korea Advanced Institute of Science and Technology, Korea Research Institute of Chemical Technology
• Original Assignee: Korea Advanced Institute of Science and Technology, Korea Research Institute of Chemical Technology
• Inventors: Lee, Sang Yup, Park, Si Jae, Lee, Seung Hwan, Song, Bong Keun, Jung, Yu Kyung, Lee, Tae Woo
• Primary Examiner(s): Monshipouri, Maryam
• Application Number:
  US14/004,437
• Publication Number:
  US 20140030775A1
• Time in Patent Office: 977 Days
• Field of Search: 435/135, 435/252.3, 435/419, 435/471
• US Class Current: 435/135
• CPC Class Codes: C12N 15/70 Vectors or expression systems specially adapted for E. coli
• CPC Class Codes: C12N 9/0006 acting on CH-OH groups as donors (1.1)
• CPC Class Codes: C12N 9/1029 transferring groups other than amino-acyl groups (2.3.1)
• CPC Class Codes: C12P 7/625 Polyesters of hydroxy carboxylic acids

Abstract

There is provided a recombinant microorganism having producibility of poly(lactate-co-glycolate) from glucose, and more particularly, a recombinant microorganism having producibility of poly(lactate-co-glycolate) without adding an exogenous glycolate precursor, and a method of preparing [poly(preparing lactate-co-glycolate)] using the same. According to the present invention, the poly(lactate-co-glycolate) in which the concentration of the glycolate fraction is high may be prepared at a high concentration without supplying exogenous glyoxylate. Therefore, the present invention may be effectively used for treatment.

Claims

1. A recombinant microorganism having producibility of poly(lactate-co-glycolate), wherein the microorganism is transfected so that a gene coding poly(hydroxyalkanoate) (PHA) synthase, a gene coding propionyl-CoA transferase, and a gene coding glycerate dehydrogenase (EC 1.1.1.26) are inserted.

2. The recombinant microorganism of claim 1, wherein the propionate CoA-transferase is propionate CoA-transferase of C. propionicum or Pct540, which is a mutant enzyme of propionate CoA-transferase of C. propionicum.

3. The recombinant microorganism of claim 1, wherein the poly(hydroxyalkanoate) (PHA) synthase is PHA synthase of Pseudomonas sp. 6-19 or a mutant enzyme of PHA synthase of Pseudomonas sp. 6-19.

4. The recombinant microorganism of claim 3, wherein the gene coding the mutant enzyme of the PHA synthase has a base sequence corresponding to an amino acid sequence including at least one mutation selected from a group consisting of E130D, S325T, L412M, S477R, S477H, S477F, S477Y, S477G, Q481M, Q481K, and Q481R in an amino acid sequence of SEQ ID No:

1.

5. The recombinant microorganism of claim 3, wherein the gene coding the mutant enzyme of the PHA synthase has a base sequence corresponding to an amino acid sequence selected from a group consisting of:

an amino acid sequence (C1335) in which E130D, S325T, L412M, S477G, and Q481M in the amino acid sequence of SEQ ID No;

1 are mutated;

an amino acid sequence (C1310) in which E130D, S477F, and Q481K in the amino acid sequence of SEQ ID No;

1 are mutated; and

an amino acid sequence (C1312) in which E130D, S477F, and Q481R in the amino acid sequence of SEQ ID No;

1 are mutated.

6. The recombinant microorganism of claim 1, wherein the gene coding glycerate dehydrogenase (EC 1.1.1.26) is derived from Pasteurella multocida.

7. The recombinant microorganism of claim 6, wherein the gene coding glycerate dehydrogenase has a base sequence of SEQ ID No:

2.

17. A method of preparing poly(lactate-co-glycolate) (PLGA) including:

culturing the recombinant microorganism of claim 1 to produce poly(lactate-co-glycolate); and

obtaining the produced PLGA.

8. A recombinant microorganism having producibility of poly(lactate-co-glycolate), wherein the microorganism is transfected so that a gene coding PHA synthase, a gene coding propionyl-CoA transferase, and a gene coding glycerate dehydrogenase (EC 1.1.1.26) are inserted, and a gene (iclR) coding isocitrate lyase regulator or aceB (malate synthase) gene is deleted.

9. The recombinant microorganism of claim 8, wherein the propionate CoA-transferase is propionate CoA-transferase of C. propionicum or Pct540, which is a mutant enzyme of propionate CoA-transferase of C. propionicum.

10. The recombinant microorganism of claim 8, wherein the poly(hydroxyalkanoate) (PHA) synthase is PHA synthase of Pseudomonas sp. 6-19 or a mutant enzyme of PHA synthase of Pseudomonas sp. 6-19.

11. The recombinant microorganism of claim 10, wherein the gene coding the mutant enzyme of the PHA synthase has a base sequence corresponding to an amino acid sequence including at least one mutation selected from a group consisting of E130D, S325T, L412M, S477R, S477H, S477F, S477Y, S477G, Q481M, Q481K, and Q481R in an amino acid sequence of SEQ ID No:

1.

12. The recombinant microorganism of claim 10, wherein the gene coding the mutant enzyme of the PHA synthase has a base sequence corresponding to an amino acid sequence selected from a group consisting of:

an amino acid sequence (C1335) in which E130D, S325T, L412M, S477G, and Q481M in the amino acid sequence of SEQ ID No;

1 are mutated;

an amino acid sequence (C1310) in which E130D, S477F, and Q481K in the amino acid sequence of SEQ ID No;

1 are mutated; and

an amino acid sequence (C1312) in which E130D, S477F, and Q481R in the amino acid sequence of SEQ ID No;

1 are mutated.

13. The recombinant microorganism of claim 10, wherein the gene coding glycerate dehydrogenase (EC 1.1.1.26) is derived from Pasteurella multocida or E. coli.

14. The recombinant microorganism of claim 13, wherein the gene coding glycerate dehydrogenase has a base sequence of SEQ ID No:

2.

15. The recombinant microorganism of claim 10, wherein both of the gene (iclR) coding isocitrate lyase regulator and a gene (aceB) coding isocitrate lyase are deleted.

16. The recombinant microorganism of claim 10, wherein the gene (aceA) coding isocitrate lyase is additionally amplified.

18. A method of preparing poly(lactate-co-glycolate) (PLGA) including:

culturing the recombinant microorganism of claim 10 to produce poly(lactate-co-glycolate); and

obtaining the produced PLGA.