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Immobilized lysine decarboxylase, its preparation, 1,5-pentanediamine preparation method and product

A lysine decarboxylase and amine-based technology, applied in the direction of immobilized enzymes, biochemical equipment and methods, fixed on/in organic carriers, etc., can solve the problem of less immobilized lysine decarboxylase, etc., To achieve the effect of solving the poor stability of use, improving the use efficiency and strong reactivity

Active Publication Date: 2021-02-19
CATHAY R&D CENT CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] At present, there are few reports on the preparation and stability of immobilized lysine decarboxylase, and there are no reports on the chemical modification of cheap carriers to make it suitable for the preparation of immobilized lysine decarboxylase

Method used

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Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0113] The preparation process of the immobilized lysine decarboxylase of the present invention can be obtained by reacting the epoxy-modified polymer carrier and the lysine decarboxylase in a buffer solution.

[0114] The amount of lysine decarboxylase relative to the epoxy-modified polymer carrier can be adjusted, and the obtained immobilized lysine decarboxylase can maintain high enzyme activity. In some embodiments, the amount of lysine decarboxylase is 1500-4000 U per gram of the epoxy-modified polymer carrier. In some embodiments, the amount of lysine decarboxylase is 2000-3500 U based on each gram of the epoxy-modified polymer carrier; in some embodiments, the amount of lysine decarboxylase is based on each gram of the ring The oxygen-modified polymer carrier is 2500-3000U.

[0115] In some embodiments, the activity concentration of the lysine decarboxylase solution can be controlled at 100-1000 U / mL.

[0116] The epoxy-modified polymer carrier and the lysine decarbox...

preparation Embodiment 1

[0133] Preparation Example 1 The preparation of lysine decarboxylase

[0134] The microbial glycerol preservation bacterium liquid that expresses lysine decarboxylase is inoculated in 500 milliliters that 100 milliliters of liquid culture medium (LB medium, comprise peptone 1%, yeast powder 0.5%, sodium chloride 1%, pH7.0) are housed Cultivate in the seed bottle at 37°C and 200rmp shaker for 15 hours to obtain seed solution. In a 5L fermenter, add 3 liters of LB medium, sterilize at 121°C for 20 minutes and then insert the above seed liquid to start fermentation (the formula of the fermentation medium is LB medium: peptone 1%, yeast powder 0.5%, sodium chloride 1 %, pH7.0), start fermentation at 30°C and 300 rpm, control the air flow rate to 0.3vvm, tank pressure to 0.04MPa, control the pH value of the fermentation broth to 7.0 during the fermentation process, and stop the fermentation after 25 hours of fermentation. After the fermentation, collect the cells (that is, wet cel...

preparation example 2

[0135] The etherification reaction of preparation example 2 cellulose acetate

[0136] (1) Add 2L of NaOH solution with a concentration of 0.5mol / L to 100g of cellulose acetate, add it into hot water, hydrolyze in a water bath for 1 hour, pour out the hydrolyzate, and wash it repeatedly with warm water until it is neutral and hydrolyzed of cellulose.

[0137] (2) Weigh SESA, add it to the sodium carbonate aqueous solution of 20g / L concentration, add sodium hydroxide to its final concentration of 0.1mol / L, SESA concentration is 100g / L, grind and dissolve, after stabilization, use filter paper Filter and take the clear liquid as etherified liquid.

[0138] (3) Weigh 50 g of the hydrolyzed cellulose acetate filaments prepared in step (1), add 300 g of the etherified solution prepared in step (2), and carry out the etherification reaction under the condition of heating in a water bath at 60° C. Add 4 mol / L sodium hydroxide solution dropwise, stir manually until the pH is 9-11, s...

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Abstract

The invention provides an immobilized lysine decarboxylase, which comprises epoxy-modified polymer carrier and lysine decarboxylase, and the lysine decarboxylase reacts with epoxy group and is fixed on the polymer carrier through covalent bond . The invention also provides a preparation method of the immobilized lysine decarboxylase, a method for preparing 1,5-pentanediamine by using the immobilized lysine decarboxylase, and the prepared 1,5-pentanediamine. The immobilized lysine decarboxylase is covalently bonded to the lysine decarboxylase through the epoxy group of the polymer carrier to form a stable immobilized lysine decarboxylase with high immobilization efficiency and high recovery rate of enzyme activity , good stability, significantly reduced carrier cost, the preparation method of 1,5-pentamethylenediamine provided by the invention effectively reduces the process cost, simplifies the separation steps of 1,5-pentamethylenediamine solution and enzyme, and increases 1 The degree of automation in the production of 5-pentanediamine has promoted the industrialization process of biological production of 1,5-pentanediamine.

Description

technical field [0001] The invention relates to the field of biochemical industry, in particular to an immobilized lysine decarboxylase, a preparation method thereof, a preparation method of 1,5-pentanediamine using the immobilized lysine decarboxylase and a product thereof. Background technique [0002] Biocatalytic reactions have the characteristics of mild reaction conditions, high selectivity, and high reaction efficiency, so they are increasingly used in the field of chemical synthesis. However, due to the poor stability of biological enzymes in in vitro industrial environments and the high cost of enzyme preparations, the practical application of biocatalysis in the industrial field is greatly limited. [0003] Immobilized enzyme technology not only effectively solves the problem of repeated use of catalysts, but also significantly improves the stability and catalytic properties of biological enzymes (substrate affinity, stereoselectivity, etc.) ). At present, the re...

Claims

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Application Information

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IPC IPC(8): C12N11/12C12N11/087C12N11/096C12N11/084C12N11/089C12N11/02C12P13/00
CPCC12N9/88C12N11/02C12N11/08C12N11/12C12P13/001C12Y401/01018
Inventor 徐冠珠于丽珺刘修才
Owner CATHAY R&D CENT CO LTD
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