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Biological preparation method of R-3-aminobutanol

An aminobutanol and biological preparation technology, applied in the directions of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., to achieve the effects of low process cost, low price, high conversion rate and product concentration

Inactive Publication Date: 2014-11-05
洛阳华荣生物技术有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, racemic 3-aminobutanol can be synthesized by chemical methods under the catalysis of ammonium salt and nickel (Chem.Abstr., 1947 , p. 6199), However, there is no report on the synthesis of biocatalyzed

Method used

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  • Biological preparation method of R-3-aminobutanol
  • Biological preparation method of R-3-aminobutanol

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preparation example Construction

[0020] A kind of biological preparation method of R-3-aminobutanol, its preparation method comprises the following steps:

[0021] Step 1, preparation of recombinant D-transaminase genetically engineered bacteria

[0022] Choose from Arthrobacter. sp. The wild sequence of D-transaminase was artificially designed, and the designed gene sequence is shown as SEQ ID NO: 1; the sequence was cloned into the Nde I and Hind III sites of pET24a through whole gene synthesis; transformed Escherichia coli DH5α state cells; after picking positive transformants and identifying them by sequencing, the recombinant expression vector was obtained; the recombinant expression vector was transformed into Escherichia coli BL21 (DE3) strain to obtain recombinant D-transaminase genetically engineered bacteria that could induce the expression of recombinant D-transaminase;

[0023] Step 2, preparation of recombinant D-transaminase

[0024] Inoculate recombinant D-transaminase genetically engineered ...

Embodiment 1

[0030] A kind of biological preparation method of R-3-aminobutanol, its preparation method comprises the following steps:

[0031] Step 1, preparation of recombinant D-transaminase genetically engineered bacteria

[0032] Choose from Arthrobacter. sp. The wild sequence of D-transaminase was artificially designed, and the designed gene sequence is shown as SEQ ID NO: 1; the sequence was cloned into the Nde I and Hind III sites of pET24a through whole gene synthesis; transformed Escherichia coli DH5α state cells; after picking positive transformants and identifying them by sequencing, the recombinant expression vector was obtained; the recombinant expression vector was transformed into Escherichia coli BL21 (DE3) strain to obtain recombinant D-transaminase genetically engineered bacteria that could induce the expression of recombinant D-transaminase;

[0033] Step 2, preparation of recombinant D-transaminase

[0034] Inoculate recombinant D-transaminase genetically engineered ...

Embodiment 2

[0040] A kind of biological preparation method of R-3-aminobutanol, its preparation method comprises the following steps:

[0041] Step 1, preparation of recombinant D-transaminase genetically engineered bacteria

[0042] Choose from Arthrobacter. sp. The wild sequence of D-transaminase was artificially designed, and the designed gene sequence is shown as SEQ ID NO: 1; the sequence was cloned into the Nde I and Hind III sites of pET24a through whole gene synthesis; transformed Escherichia coli DH5α state cells; after picking positive transformants and identifying them by sequencing, the recombinant expression vector was obtained; the recombinant expression vector was transformed into Escherichia coli BL21 (DE3) strain to obtain recombinant D-transaminase genetically engineered bacteria that could induce the expression of recombinant D-transaminase;

[0043] Step 2, preparation of recombinant D-transaminase

[0044] Inoculate recombinant D-transaminase genetically engineered ...

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Abstract

The invention provides a biological preparation method of R-3-aminobutanol. The method includes: conducting whole-gene synthesis according to an artificially designed sequence shown as SEQ ID NO:1, and performing cloning into pET24a so as to obtain a recombinant expression vector; transferring the recombinant expression vector into E.coli to obtain a recombinant D-transaminase genetic engineering bacterium; culturing the bacterium to prepare recombinant D-transaminase; adding 3-carbonyl butanol with a final concentration of 10-300mmol / L, dimethyl sulfoxide or acetonitrile with a mass percentage concentration of 2-15%, pyridoxal phosphate with a final concentration of 0.1-1mmol / L, isopropylamine or D-alanine with a final concentration of 0.02-1mol / L, and recombinant D-transaminase with a mass percentage concentration of 0.01-1% are added into the reaction solution to compose a reaction system for reaction; and at the end of the reaction, extracting R-3-aminobutanol from the reaction solution. With a low cost, the method provided by the invention can achieve a conversion rate of more than 97% and a product yield of greater than 87%, and does not generate by-product, thus being more suitable for industrial application.

Description

technical field [0001] The present invention relates to a kind of R-3-aminobutanol, specifically a kind of biological preparation 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] At present, racemic 3-aminobutanol can be synthesized by chemical methods under the catalysis of ammonium salt and nickel (Chem.Abstr., 1947, p. 6199), but there is no report on the ...

Claims

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Application Information

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IPC IPC(8): C12P13/00C12N15/70C12N1/21C12N9/10C12R1/19
Inventor 范文超丁鹏
Owner 洛阳华荣生物技术有限公司
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