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A kind of preparation method and application of carotenoid 9, 10' dioxygenase

A carotene and dioxygenase technology, applied in the field of bioengineering, can solve the problems of low yield, high requirement of reaction conditions, long reaction time of β-ionone, etc., and achieves short reaction time, less reaction products and optimized Effects of reaction conditions

Active Publication Date: 2019-09-13
ZHENGZHOU UNIVERSITY OF LIGHT INDUSTRY
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Problems solved by technology

[0004] The invention provides a preparation method of carotenoid 9, 10' dioxygenase, which solves the problems of long reaction time, low yield and high requirements on reaction conditions in the industrial production of β-ionone

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  • A kind of preparation method and application of carotenoid 9, 10' dioxygenase
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  • A kind of preparation method and application of carotenoid 9, 10' dioxygenase

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Embodiment Construction

[0030] 1. Preparation of Enterobacter carotenoid 9, 10' dioxygenase

[0031] The preparation method of Enterobacteriaceae carotenoid 9, 10' dioxygenase, including strain, plasmid, enzyme and culture medium, primer design, PCR amplification and construction of recombinant plasmid, induced expression of gene and purification of protein. Specific steps are as follows:

[0032] (1) Strain, plasmid, enzyme and culture medium:

[0033] Escherichia coli ( Escherichia coli )DH5α, restriction endonuclease Nde I and Sal I. Pyrobest DNA polymerase, DNA ligation kit ver.2.1, bacterial genome DNA extraction kit ver.3.0 were purchased from TAKARA company; Escherichia coli Escherichia coli BL21-codonPlus(DE3)-RIL was purchased from Novagen’s LB medium: each liter contained 10 g of Tryptone, 5 g of Yeast extract, and 10 g of NaCl; the concentration of ampicillin was 100 mg / L, and the concentration of chloramphenicol was 34 mg / L ; Enterobacter hallii ( Enterobacter hormaechei ) is...

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Abstract

The invention belongs to the field of bioengineering and relates to a preparation method and application of a carotenoid 9,10' dioxygenase. The preparation method comprises the following steps: cultivating enterobacter hormaechei, and extracting a whole genome sequence; constructing a recombinant plasmid of a carotenoid 9,10' dioxygenase gene; inducing expression of the carotenoid 9,10'dioxygenase gene, and purifying zymoprotein; and analyzing and identifying enzymatic properties of the carotenoid 9,10' dioxygenase. The prepared carotenoid 9,10' dioxygenase gene can efficiently catalyze and synthesize beta-ionone, catalysis specificity is high, reaction products are few, and reaction time is short; a novel enzyme resource different from discovered carotenoid 9,10' dioxygenase is obtained; the carotenoid 9,10' dioxygenase has different amino acid sequences and enzymatic characteristics; and reaction conditions are optimized, and yield of the beta-ionone reaches 79.4%.

Description

technical field [0001] The invention belongs to the field of bioengineering, and relates to a preparation method and application of carotenoid 9, 10' dioxygenase. Background technique [0002] β-ionone is a natural fragrance and an important intermediate, widely used in food, cosmetics and pharmaceutical industries. As a representative of cyclized isoprenoids, β-ionone has a wide range of biological activities, especially exhibiting good antitumor activity. As a plant secondary metabolite, β-ionone is extracted from plant essential oils in a small amount and is expensive, and currently it is mainly synthesized by chemical methods. The chemical method has high requirements on reaction conditions and separation equipment, the product is not easy to separate, and the waste liquid produced pollutes the environment. In addition, with the improvement of living standards and the enhancement of health awareness, the safety of chemically synthesized β-ionone has been questioned mor...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/02C12N15/53C12N15/70C12P7/26
Inventor 魏涛毛多斌黄申白冰臧杰杨昆鹏江森李付良
Owner ZHENGZHOU UNIVERSITY OF LIGHT INDUSTRY
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