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a kind of adding fe 2+ Improve the method for Escherichia coli engineering bacteria to synthesize 5-aminolevulinic acid

A technology of aminolevulinic acid and Escherichia coli, applied in the fields of metabolic engineering and microbial fermentation, can solve problems such as high cost, unsuitability for large-scale industrial production, etc., and achieve the effect of increasing yield

Active Publication Date: 2018-01-16
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the particularity of photosynthetic bacteria, their cost is high and they are not suitable for large-scale industrial production

Method used

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  • a kind of adding fe <sup>2+</sup> Improve the method for Escherichia coli engineering bacteria to synthesize 5-aminolevulinic acid
  • a kind of adding fe <sup>2+</sup> Improve the method for Escherichia coli engineering bacteria to synthesize 5-aminolevulinic acid
  • a kind of adding fe <sup>2+</sup> Improve the method for Escherichia coli engineering bacteria to synthesize 5-aminolevulinic acid

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 adds Fe 2+ Effects on engineered strains of Escherichia coli

[0039] Strain: LADF-6:E.coli BL21(DE3) / pRSFDuet-1-hemA-hemL-hemF pETDuet-1-hemD. Add 2.5mg / L of FeSO at 0h of fermentation 4 ·7H 2 0, analyze the growth situation of cell and the accumulation situation of target product ALA, the result is as follows figure 1 As shown, adding Fe 2+ Afterwards, the cell mass increased, and the synthesis of the product ALA was also accelerated.

Embodiment 2

[0040] Embodiment 2 optimizes Fe 2+ Concentration Boosts ALA Production

[0041] Strain: LADF-6:E.coli BL21(DE3)pRSFDuet-1-hemLA-hemFpETDuet-1-hemD.

[0042] Analysis and investigation of recombinant Escherichia coli in the addition of different concentrations of Fe 2+ When the accumulation of ALA, the results are as follows figure 2 As shown, adding different concentrations of Fe 2+ The amount of ALA accumulated by recombinant Escherichia coli was different when the Fe 2+ With the increase of the addition amount, the ALA production gradually increased, but when it was greater than 10mg / L, the ALA accumulation decreased.

Embodiment 3

[0043] Embodiment 3 Recombinant bacteria 3L fermenter fermentation verification

[0044] Strain: LADF-6:E.coli BL21(DE3)pRSFDuet-1-hemLA-hemFpETDuet-1-hemD.

[0045] Recombinant Escherichia coli LADF-6 is fermented and produced in a 3L fermenter, the inoculum size is 2%, the initial glucose concentration is 33g / L, and 1.0-15mg / L FeSO is added at 0h 4 , 0.1-0.5mM IPTG induction and corresponding antibiotics, ALA began to accumulate in large quantities after 10h, and the output was the highest at about 30h, which was 3.85g / L ( image 3 ).

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Abstract

The invention discloses a method for enhancing engineered escherichia coli synthesized 5-aminolevulinic acid by adding Fe<2+> and belongs to the field of metabolic engineering and microbial fermentation. By using established escherichia coli BL21(DE3)LADF-6 for synthesizing 5-aminolevulinic acid as an original strain, the influence of adding Fe<2+> into a culture medium to synthesis ALA is inspected. Verified by fermentation, the target product ALA output is obviously improved and the ALA output after 30 hours is 2.25g / L. On this basis, by optimizing the initial addition of Fe<2+> and carrying out scale-up culture in a 3L fermentation tank, when 10mg / L of Fe<2+> is added, the ALA output is 3.85g / L.

Description

technical field [0001] The present invention relates to a kind of adding Fe 2+ The invention discloses a method for improving the synthesis of 5-aminolevulinic acid by Escherichia coli engineering bacteria, belonging to the fields of metabolic engineering and microbial fermentation. Background technique [0002] 5-aminolevulinic acid (5-aminolevulinic acid, ALA), the molecular formula is C 5 o 3 NH 9 , with a molecular weight of 131.13 and a melting point of 149-151°C, it is the biosynthesis of chlorophyll, heme, and vitamin B 12 and other key precursors. As a safe, selective, and permeable photodynamic drug, ALA has gradually attracted attention in the medical field, and has been successfully used in the diagnosis and photodynamic therapy of skin cancer, bladder cancer, digestive tract cancer, and lung cancer. In addition, because ALA is degradable in nature, it is also widely used in the field of pesticides, such as a new type of pollution-free photoactivated pesticid...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P13/00C12R1/19
Inventor 康振陈坚堵国成张俊丽
Owner JIANGNAN UNIV
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