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Copolymer comprising 4-hydroxybutyrate unit and lactate unit and its manufacturing method

A technology of lactate terpolymer and hydroxybutyrate, which is applied in DNA/RNA fragments, recombinant DNA technology, fermentation, etc., can solve the problem of complex preparation methods of high-molecular-weight PLA and the difficulty of removing solvents or chain coupling agents , did not disclose the preparation of copolymers and other issues

Active Publication Date: 2009-10-07
LG CHEM LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it has the following disadvantages: as the preparation method of high molecular weight PLA is complicated due to the addition of solvent or chain coupling agent, and it is not easy to remove the solvent or chain coupling agent
However, this publication does not disclose a method for preparing copolymers using lactoyl-CoA and 3-hydroxyalkanoate-CoA

Method used

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  • Copolymer comprising 4-hydroxybutyrate unit and lactate unit and its manufacturing method
  • Copolymer comprising 4-hydroxybutyrate unit and lactate unit and its manufacturing method
  • Copolymer comprising 4-hydroxybutyrate unit and lactate unit and its manufacturing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1: Construction of a recombinant plasmid comprising pct gene and PHA synthase gene.

[0041] Recombinant plasmids pPs619C1300-CPPCT and pTacCpPctNCvEC containing pct gene and PHA synthase gene were constructed to prepare copolymers containing 4-hydroxybutyrate unit and lactate unit.

[0042] (1) Construction of plasmid pPs619C1300-CPPCT

[0043] The propionyl-CoA transferase (CP-PCT) gene derived from Clostridium propionicum was used as the pct gene, and the PHA synthase gene derived from Pseudomonas 6-19 was used as the PHA synthase gene .

[0044] Such as figure 1 An operon for a constitutive expression system expressing PHA synthase and CP-PCT together was constructed. CP-PCT is known to be toxic to host microorganisms. In other words, in the tac promoter or T7 promoter expression system induced by IPTG, which is widely used for the expression of recombinant proteins, all microorganisms die within a short time after the addition of the inducer. For this ...

Embodiment 2

[0083] Embodiment 2: Preparation of 4-hydroxybutyrate-lactate copolymer

[0084] E. coli Top 10 (Invitrogen) was transformed with pPs619C1300-CPPCT obtained in Example 1 together with pMCSPtbBuk to obtain E. coli Top 10 / pPs619C1300-CPPCT / pMCSPtbBuk.

[0085] The 4-hydroxybutyrate-lactate copolymer was obtained by cultivating the transformant in the following two steps: firstly, the transformed recombinant E. Vegetable LB medium (Bacto TM Tryptone (BD) 10g / L, Bacto TM After culturing in yeast extract (BD) 5 g / L, NaCl (amresco) 10 g / L) for 24 hours, the medium was centrifuged at 4° C. at 1000 g for 15 minutes to collect cells.

[0086] The collected cells were cultured in MR medium (glucose 10 g, KH 2 PO 4 6.67g, (NH 4 ) 2 HPO 4 4g, MgSO 4 ·7H 2 O 0.8g, citric acid 0.8g and trace metal solution 5mL; trace metal solution composition: 5M HCl 5mL per L, FeSO 4 ·7H 2 O 10g, CaCl 2 2g, ZnSO 4 ·7H 2 O 2.2g, MnSO 4 4H 2 O 0.5g, CuSO 4 ·5H 2 O 1g, (NH 4 ) 6 Mo 7 ...

Embodiment 3

[0089] Embodiment 3: Preparation of 4-hydroxybutyrate-3-hydroxypropionate ester terpolymer

[0090] In addition to MR medium containing 2g / L 4-hydroxybutyrate (4-HB), 2g / L 3-hydroxypropionate (3-HP), 100mg / L ampicillin and 30mg / L chloramphenicol Instead of anaerobically culturing the collected cells in MR medium containing 2g / L 4-HB, 100mg / L ampicillin and 30mg / L chloramphenicol for 3 days, prepare 4-hydroxybutyrate according to the method of Example 2 - 3-Hydroxypropionate-lactate terpolymer.

[0091] As a result of the analysis, methyl-4-hydroxybutyrate, methyl-3-hydroxypropionate, and methyl-lactate were detected in the E. coli Top10 / pPs619C1300-CPPCT / pMCSPtbBuk transformant, which means that by Preparation of Novel 4-Hydroxybutyrate-3-Hydroxypropionate-Lactate Terpolymers [Poly(4-Hydroxybutyrate-Co-3-Hydroxypropionate-Co-Lactate) acid ester)]. The obtained 4-hydroxybutyrate-3-hydroxypropionate-lactate terpolymer 1 H-NMR and GC-MSD results are shown in Image 6 with 7...

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Abstract

The present invention relates to a copolymer comprising 4-hydroxybutyrate monomer unit and lactate monomer unit, a copolymer 4-hydroxybutyrate monomer unit, lactate monomer unit and 3-hydroxyalkanoate, or their preparing method. More specifically, the present invention relates to a method for preparing a copolymer comprising lactate monomer; 4-hydroxybutyrate monomer; and optionally 3-hydroxyalkanoate, wherein the method comprises culturing a cell or plant comprising the gene of enzyme converting lactate and 3-hydroxyalkanoate into lactyl-CoA and 3-hydroxyalkanoyl-CoA, respectively, phosphotransbutylase gene, butyrate kinase gene and polyhydroxyalkanoate synthase gene together, and the copolymer made by the method. The copolymer of the present invention is a biodegradable polymer being able to be usefully used instead of conventional synthetic plastic, and the copolymer can be used for medical use.

Description

technical field [0001] The present invention relates to a copolymer comprising 4-hydroxybutyrate monomer units and lactate monomer units or comprising 4-hydroxybutyrate monomer units, lactate monomer units and 3-hydroxyalkanoate monomer units Copolymers of monomeric units and processes for the preparation of such polymers. Background technique [0002] Polylactate (PLA) is a typical biodegradable polymer derived from lactate, which has various applications as a general or medical polymer. Currently, PLA is prepared by polymerizing lactic acid esters produced by microbial fermentation, but only low molecular weight (1000-5000 Daltons) PLA can be produced by direct polymerization of lactic acid esters. For the synthesis of high molecular weight (>100,000 Daltons) PLA, a method of polymerizing low molecular weight PLA obtained by direct polymerization of lactate via a chain coupling agent can be used. However, it has disadvantages such as complicating the preparation metho...

Claims

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Application Information

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IPC IPC(8): C08G63/91C08G63/08
CPCC08G63/00C08G63/06C08G63/08C08G63/78C08L67/00C08L2203/02C12N15/52C12P7/56
Inventor 朴时载李相贤李恩政姜惠玉金泰完梁宅镐李相烨
Owner LG CHEM LTD
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