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Method for producing R-3-aminobutanol

A technology of aminobutanol and its production method, which is applied in the direction of biochemical equipment and methods, enzymes, and the use of carriers to introduce foreign genetic material, etc., can solve the problems of no biocatalysis, etc., and achieve the effects of easy separation, reduced separation cost, and low price

Active Publication Date: 2014-12-03
洛阳华荣生物技术有限公司
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, by chemical method, racemic 3-aminobutanol (Chem.Abstr., can be synthesized under the catalysis of ammonium salt and nickel 1947 , p. 6199), but there is no report on biocatalysis, especially synthesis by amino acid dehydrogenase

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  • Method for producing R-3-aminobutanol
  • Method for producing R-3-aminobutanol

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preparation example Construction

[0017] A kind of production method of R-3-aminobutanol, its preparation method comprises the following steps:

[0018] Step 1, preparation of recombinant D-amino acid dehydrogenase genetically engineered bacteria

[0019] Choose from Corynebacterium glutamicum The D-amino acid dehydrogenase gene sequence of ATCC13032 was artificially designed, and the designed gene sequence is shown in SEQ ID NO: 1; the sequence was cloned into the expression vector pET22b by Nde I and Hind III enzymes through whole gene synthesis site, transform Escherichia coli DH5α competent cells; pick positive transformants and sequence and identify them, and obtain recombinant expression vectors; transfer recombinant expression vectors into Escherichia coli BL21 (DE3) strains, and obtain recombinant D-amino acid desensitizers that can induce expression Hydrogenase recombinant D-amino acid dehydrogenase genetically engineered bacteria;

[0020] Step 2, preparation of recombinant D-amino acid dehydroge...

Embodiment 1

[0026] A kind of production method of R-3-aminobutanol, its preparation method comprises the following steps:

[0027] Step 1, preparation of recombinant D-amino acid dehydrogenase genetically engineered bacteria

[0028] Choose from Corynebacterium glutamicum The D-amino acid dehydrogenase gene sequence of ATCC13032 was artificially designed, and the designed gene sequence is shown in SEQ ID NO: 1; the sequence was cloned into the expression vector pET22b by Nde I and Hind III enzymes through whole gene synthesis site, transform Escherichia coli DH5α competent cells; pick positive transformants and sequence and identify them to obtain recombinant expression vectors; transfer the recombinant expression vectors into Escherichia coli BL21 (DE3) strains to obtain recombinant D-amino acid desensitizers that can induce expression Hydrogenase recombinant D-amino acid dehydrogenase genetically engineered bacteria;

[0029] Step 2, preparation of recombinant D-amino acid dehydroge...

Embodiment 2

[0035] A kind of production method of R-3-aminobutanol, its preparation method comprises the following steps:

[0036] Step 1, preparation of recombinant D-amino acid dehydrogenase genetically engineered bacteria

[0037] Choose from Corynebacterium glutamicum The D-amino acid dehydrogenase gene sequence of ATCC13032 was artificially designed, and the designed gene sequence is shown in SEQ ID NO: 1; the sequence was cloned into the expression vector pET22b by Nde I and Hind III enzymes through whole gene synthesis site, transform Escherichia coli DH5α competent cells; pick positive transformants and sequence and identify them, and obtain recombinant expression vectors; transfer recombinant expression vectors into Escherichia coli BL21 (DE3) strains, and obtain recombinant D-amino acid desensitizers that can induce expression Hydrogenase recombinant D-amino acid dehydrogenase genetically engineered bacteria;

[0038] Step 2, preparation of recombinant D-amino acid dehydroge...

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Abstract

The invention relates to a method for producing R-3-aminobutanol. The method comprises the following steps: artificially designing a sequence shown in SEQID NO: 1, carrying out full gene synthesis, cloning the synthesized gene fragments into an expression vector pET22b to prepare a recombinant expression vector, transferring the recombinant expression vector into Escherichia coli to prepare a genetically engineered bacterium of a recombinant D-amino acid dehydrogenase, culturing the genetically engineered bacterium to prepare the recombinant D-amino acid dehydrogenase, sequentially adding a substrate of which the final concentration is 10-300mmol / L, a 2-15wt% cosolvent, a cofactor of which the final concentration is 0.1-1mmol / L, an amino donor of which the final concentration is 0.02-1mmol / L and the 0.01-1wt% D-amino acid dehydrogenase into a reaction solution to constitute a reaction system, reacting, after the reaction is completed, and extracting R-3-aminobutanol in the reaction solution. The method disclosed by the invention has the advantages that the cost is low, the conversion rate is greater than 95%, the product yield is greater than 85%, no by-product is generated and method is more suitable for industrial applications.

Description

technical field [0001] The invention belongs to the field of biocatalysis and relates to a production method of R-3-aminobutanol. Background technique [0002] Biocatalysis has the characteristics of mild reaction conditions, high efficiency, strong stereo and regioselectivity, and environmental friendliness, so biocatalysis has been widely used in the research and development of new drugs. In addition, the use of rational design and directed evolution to modify and optimize biocatalysts, such as improving catalyst stability and substrate selectivity, will enable them to be better applied to production processes. [0003] R-3-aminobutanol is widely used in organic synthesis and pharmaceutical production, and its structural formula is as follows: [0004] [0005] Currently using chemical methods, under the catalysis of ammonium salt and nickel, racemic 3-aminobutanol can be synthesized ( Chem. Abstr., 1947 , p. 6199), but there is no report on the synthesis by bioc...

Claims

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Application Information

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IPC IPC(8): C12P13/00C12N15/70C12N1/21C12N9/06
Inventor 范文超丁鹏
Owner 洛阳华荣生物技术有限公司
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